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Biomedical subjects

C Li

Publications and source records attributed to C Li.

At least 19 recordsLinked to original sources
PubMed1999

Tenascin-C is expressed in macrophage-rich human coronary atherosclerotic plaque.

BACKGROUND: Tenascin is a large extracellular matrix glycoprotein generally found in adult tissues undergoing active remodeling such as healing wounds and tumors. To determine the potential role of tenascin-C (TN-C) in the pathophysiology of atherosclerosis, we investigated the pattern of expression of TN-C in human coronary atherosclerotic plaques. METHODS AND RESULTS: Immunohistochemical staining and in situ hybridization demonstrated minimal and random expression of TN-C in fibrotic but lipid-poor atherosclerotic plaques. In contrast, all plaques with an organized lipid core or ruptured intimal surface strongly expressed TN-C, which was preferentially concentrated around the lipid core, shoulder regions, and ruptured area of the plaques but not in the fibrous cap. TN-C was not detected in normal arterial tissue. To identify the cellular source of TN-C, the plaques were stained with smooth muscle cell- and macrophage-specific antibodies. TN-C expression correlated with the infiltration of macrophages. Northern blot and immunoprecipitation analysis showed that macrophages expressed 7. 0-kb TN-C mRNA and 220-kDa protein. Reverse transcription-polymerase chain reaction of total RNA derived from macrophages showed that they express the small isoform of TN-C. Zymogram analysis revealed that macrophages markedly increased MMP-9 expression. CONCLUSIONS: This study demonstrates that the level of TN-C expression correlates with the degree of inflammation present, not with plaque size. In addition, cultured macrophages have the capacity to express the TN-C gene. These findings suggest the significance of macrophages in the remodeling of atherosclerotic plaque matrix composition.

Adult↗
PubMed1999

Induction of low density lipoprotein receptor (LDLR) transcription by oncostatin M is mediated by the extracellular signal-regulated kinase signaling pathway and the repeat 3 element of the LDLR promoter.

Oncostatin M (OM) activates the transcription of the human low density lipoprotein receptor (LDLR) in HepG2 cells through a sterol-independent mechanism. Our previous studies showed that mutations within the repeat 3 sequence of the LDLR promoter significantly decreased OM activity on LDLR promoter luciferase reporter constructs that contain the sterol responsive element-1 (repeat 2) and Sp1 binding sites (repeats 1 and 3). In this study, we investigated the signal transduction pathways that are involved in OM-induced LDLR transcription. In HepG2 cells, OM induced a rapid increase in LDLR mRNA expression, with increases detected at 30 min and maximal induction at 1 h. This OM effect was not blocked by protein synthesis inhibitors, inhibitors of p38 kinase, phosphatidylinositol 3-kinase, or c-Jun N-terminal kinase, but OM activity was completely abolished by pretreating cells with inhibitors of the extracellular signal-regulated kinase (ERK) kinase (mitogen/ERK kinase (MEK)). To investigate whether the repeat 3 sequence of the LDLR promoter is the OM-responsive element that converts ERK activation at the promoter level, three luciferase reporters, pLDLR-TATA containing only the TATA-like elements of the promoter, pLDLR-R3 containing repeat 3 and the TATA-like elements, and pLDLR-234 containing repeats 1, 2, 3 and the TATA-like elements were constructed and transiently transfected into HepG2 cells. OM had no effect on the basal promoter construct pLDLR-TATA; however, including a single copy of repeat 3 sequence in the TATA vector (pLDLR-R3) resulted in a full OM response. The activity of OM on pLDLR-R3 was identical to that of pLDLR-234. Importantly, the ability of OM to increase luciferase activities in both pLDLR-R3- and pLDLR-234-transfected cells was blocked in a dose-dependent manner by inhibition of MEK. These results demonstrate that the mitogen-activated protein kinase MEK/ERK cascade is the essential signaling pathway by which OM activates LDLR gene transcription and provide the first evidence that the repeat 3 element is a new downstream target of ERK activation.

Antineoplastic Agents↗
PubMed1999

Breast carcinoma: vascular density determined using CD105 antibody correlates with tumor prognosis.

Angiogenesis is essential for tumor growth and metastasis. There are conflicting reports as to whether microvessel density (IMD) in breast cancers is associated with prognosis. This could be due to the use of different antibodies to endothelial cell markers, variation in tissue pretreatment protocols, and nonstandardized counting methods. We have assessed the IMD in 106 breast carcinomas using a pan-endothelial marker, CD34, and a recently described mAb to CD105, which preferentially reacts with endothelial cell in angiogenic tissues. IMD values (separated as above or below median) for CD105 expression showed a statistically significant correlation with overall (P = 0.0029) and disease-free survival (P = 0.0362). In contrast, blood vessel counts using a panendothelial marker CD34 did not correlate with overall or disease-free survival (P = 0.2912 and P = 03153, respectively). When IMD values were subdivided into quartiles and assessed for their prognostic values, there was a statistically significant difference in the overall survival across CD105, but not CD34, values (P = 0.0017 and P = 0.7997, respectively) and also disease-free survival (P = 0.0431 and P = 0.5066, respectively). Further analysis of IMD values demonstrated that there were no deaths in the lowest quartile for CD105 and it differed from the other three quartiles. However, examination of clinical details of patients in the lowest quartile failed to reveal clustering of patients known to be associated with low-risk factors. Multivariate analysis confirmed that IMD values using CD105 were an independent prognostic factor. These results suggest that the ability to quantitatively distinguish between tumor neovascularization and preexisting vessels may be important in the assessment of tumor angiogenesis, but requires confirmation in a greater number of patients with a longer follow-up.

Adult↗
PubMed1999

Control of sexually transmitted diseases for AIDS prevention in Uganda: a randomised community trial. Rakai Project Study Group.

BACKGROUND: The study tested the hypothesis that community-level control of sexually transmitted disease (STD) would result in lower incidence of HIV-1 infection in comparison with control communities. METHODS: This randomised, controlled, single-masked, community-based trial of intensive STD control, via home-based mass antibiotic treatment, took place in Rakai District, Uganda. Ten community clusters were randomly assigned to intervention or control groups. All consenting residents aged 15-59 years were enrolled; visited in the home every 10 months; interviewed; asked to provide biological samples for assessment of HIV-1 infection and STDs; and were provided with mass treatment (azithromycin, ciprofloxacin, metronidazole in the intervention group, vitamins/anthelmintic drug in the control). Intention-to-treat analyses used multivariate, paired, cluster-adjusted rate ratios. FINDINGS: The baseline prevalence of HIV-1 infection was 15.9%. 6602 HIV-1-negative individuals were enrolled in the intervention group and 6124 in the control group. 75.0% of intervention-group and 72.6% of control-group participants provided at least one follow-up sample for HIV-1 testing. At enrolment, the two treatment groups were similar in STD prevalence rates. At 20-month follow-up, the prevalences of syphilis (352/6238 [5.6%]) vs 359/5284 [6.8%]; rate ratio 0.80 [95% CI 0.71-0.89]) and trichomoniasis (182/1968 [9.3%] vs 261/1815 [14.4%]; rate ratio 0.59 [0.38-0.91]) were significantly lower in the intervention group than in the control group. The incidence of HIV-1 infection was 1.5 per 100 person-years in both groups (rate ratio 0.97 [0.81-1.16]). In pregnant women, the follow-up prevalences of trichomoniasis, bacterial vaginosis, gonorrhoea, and chlamydia infection were significantly lower in the intervention group than in the control group. No effect of the intervention on incidence of HIV-1 infection was observed in pregnant women or in stratified analyses. INTERPRETATION: We observed no effect of the STD intervention on the incidence of HIV-1 infection. In the Rakai population, a substantial proportion of HIV-1 acquisition appears to occur independently of treatable STD cofactors.

Acquired Immunodeficiency Syndrome↗
PubMed1999

Requirement of Cdk2-cyclin E activity for repeated centrosome reproduction in Xenopus egg extracts.

The abnormally high number of centrosomes found in many human tumor cells can lead directly to aneuploidy and genomic instability through the formation of multipolar mitotic spindles. To facilitate investigation of the mechanisms that control centrosome reproduction, a frog egg extract arrested in S phase of the cell cycle that supported repeated assembly of daughter centrosomes was developed. Multiple rounds of centrosome reproduction were blocked by selective inactivation of cyclin-dependent kinase 2-cyclin E (Cdk2-E) and were restored by addition of purified Cdk2-E. Confocal immunomicroscopy revealed that cyclin E was localized at the centrosome. These results demonstrate that Cdk2-E activity is required for centrosome duplication during S phase and suggest a mechanism that could coordinate centrosome reproduction with cycles of DNA synthesis and mitosis.

Animals↗
PubMed1999

Walker mutations reveal loose relationship between catalytic and channel-gating activities of purified CFTR (cystic fibrosis transmembrane conductance regulator).

The cystic fibrosis transmembrane conductance regulator (CFTR) functions as an ATPase and as a chloride channel. It has been hypothesized, on the basis of electrophysiological findings, that the catalytic activity of CFTR is tightly coupled to the opening and closing of the channel gate. In the present study, to determine the structural basis for the ATPase activity of CFTR, we assessed the effect of mutations within the "Walker A" consensus motifs on ATP hydrolysis by the purified, intact protein. Mutation of the lysine residue in the "Walker A" motif of either the first nucleotide binding fold (CFTRK464A) or the second nucleotide binding fold (CFTRK1250A) inhibited the ATPase activity of the purified intact CFTR protein significantly, by greater than 50%. This finding suggests that the two nucleotide binding folds of CFTR are functioning cooperatively in catalysis. However, the rate of channel gating was only significantly inhibited in one of these purified mutants, CFTRK1250A, suggesting that ATPase activity may not be tightly coupled to channel gating as previously hypothesized.

Adenosine Triphosphatases↗
PubMed1999

ClC-2 activation modulates regulatory volume decrease.

ClC-2 belongs to a large family of chloride channels and its expression in certain cell types is associated with the appearance of swelling-activated chloride (Cl-) currents. In the present report, we examined the hypothesis that ClC-2 plays a role in regulatory volume decrease by expressing ClC-2 in Sf9 cells using the baculovirus system. First, we showed that ClC-2 protein expression is associated with appearance of a Cl- conductance which is activated by hypo-osmotic shock and can be distinguished from swelling-activated chloride currents endogenous to Sf9 cells on the basis of its pharmacology and specific inhibition by an anti-ClC-2 antibody. Second, we show that the rate of regulatory volume decrease is significantly enhanced in Sf9 cells expressing ClC-2 protein. Hence, our data support the hypothesis that ClC-2 is capable of mediating regulatory volume decrease.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1999

Isolation, pharmacology and gene organization of KPSFVRFamide: a neuropeptide from Caenorhabditis elegans.

To date, 53 peptides with C-terminal RFamides have been identified by the genome sequencing project in the nematode, Caenorhabditis elegans. In this study the FMRFamide-related peptide (FaRP) KPSFVRFamide (879.90 Da [MH]+) was structurally characterized from extracts of the nematode, Caenorhabditis elegans. Two copies of KPSFVRFamide are encoded by a gene designated flp-9. RT-PCR identified a single cDNA product which was confirmed as flp-9 by sequence determination. Flp-9 cDNA was isolated from larval stages of C. elegans but was not detected in adult worms, indicating that its expression is may be developmentally regulated. KPSFVRFamide displays sequence homology to the nematode peptide, KPNFIRFamide (PF4). The physiological effects of KPSFVRFamide, PF4 and the chimeras, KPNFVRFamide and KPSFIRFamide, were measured on body wall muscle and the vagina vera of the parasitic nematode, Ascaris suum. KPNFVRFamide and KPNFIRFamide had Cl--dependent inhibitory activity on innervated and denervated muscle-preparations, whereas KPSFVRFamide and KPSFIRFamide did not elicit a detectable physiological effect. Although all 4 peptides had inhibitory effects on the vagina vera, KPSFVRFamide and KPSFIRFamide (threshold, >/=0.1 microM) were less potent than KPNFVRFamide and KPNFIRFamide (threshold, >/=10 nM).

Amino Acid Sequence↗
PubMed1999

Allosteric regulation of even-skipped repression activity by phosphorylation.

The Drosophila homeodomain protein Even-skipped (Eve) is a well characterized transcriptional repressor. Here, we show that Eve's ability to function in vitro is negatively regulated by phosphorylation. DNA-binding activity was unaffected by phosphorylation, but phosphorylated Eve was unable to interact with the TATA-binding protein (TBP), a known target for repression. Unexpectedly, phosphorylation of the Eve N terminus, which is dispensable for repression and TBP binding, was necessary and sufficient to inactivate Eve. LiCl, which specifically inhibits glycogen synthase kinase-3 (GSK-3), reduced Eve phosphorylation in nuclear extract and blocked inhibition of repression. In addition, Eve was phosphorylated and inactivated by purified GSK-3 beta plus casein kinase II. Our results suggest a novel mechanism of transcriptional control involving phosphorylation-induced allosteric interference with a repressive protein-protein interaction.

Allosteric Regulation↗
PubMed1999

Two diterpenoids from the roots of gaultheria yunnanensis

Two new diterpenoids, gaultheric acid (1) and gaultheronoterpene (2), were isolated from the roots ofGaultheria yunnanensis. Their structures were elucidated as 12-hydroxy-13-acetyl-8,11, 13-podocarpatrien-18-oic acid (1) and 3beta, 12-dihydroxy-13-acetyl-4(18),8,11,13-podocarpatetraene (2) on the basis of spectral analysis.

Journal Article↗
PubMed1999

An improved alkaline lysis method for minipreparation of plasmid DNA.

This study is to improve the digestion pattern of miniprepped plasmid analyzed on gel. Frequently, some ambiguous DNA bands, which are suspected to be denatured DNA molecules, appear during electrophoresis of enzyme digested miniprepped plasmids. By employing Southern hybridization of two identical gels, one had been treated with denaturation-neutralization step and another without such treatment, we confirmed that many of these ambiguous DNA bands were single-stranded (SS) DNA molecules. The presence of SS DNA was due to the use of excess amount of NaOH during plasmid DNA purification with the conventional alkaline lysis method. We, therefore, modified the procedure and recommend that a half amount of NaOH (0.1N instead of 0.2N) should be used when isolating small quantity of plasmid DNA with the method.

Electrophoresis, Agar Gel↗
PubMed1999

A Lys644Glu substitution in fibroblast growth factor receptor 3 (FGFR3) causes dwarfism in mice by activation of STATs and ink4 cell cycle inhibitors.

Missense mutations of human fibroblast growth factor receptor 3 (FGFR3) result in several skeletal dysplasias, including hypochondroplasia, achondroplasia and thanatophoric dysplasia. To study the function of FGFR3 in bone growth and to create animal models for the FGFR3-related inherited skeletal disorders, we introduced a point mutation (Lys644Glu) into the murine FGFR3 genome using a knock-in approach. We found that the Lys644Glu mutation resulted in retarded endochondral bone growth with severity directly linked to the expression level of the mutated Fgfr3. Mice heterozygous for the mutation ( Fgfr3(TD/+) ) expressed the mutant allele at approximately 20% of the wild-type level and exhibited a mild bone dysplasia. However, when the copy number of the mutant allele increased from one (Fgfr3(TD/+) to two (Fgfr3(TD/TD), the retardation of bone growth became more severe and showed phenotypes resembling those of achondroplasia patients, characterized by a dramatically reduced proliferation of growth plate cartilage, macrocephaly and shortening of the long bones, which was most pronounced in the femur. Molecular analysis revealed that expression of the mutant receptor caused the activation of Stat1, Stat5a and Stat5b, and the up-regulation of p16, p18 and p19 cell cycle inhibitors, leading to dramatic expansion of the resting zone of chondrocytes at the expense of the proliferating chondrocytes. The mutant growth plates consequently were in a less active state and generated fewer maturing and hypertrophic chondrocytes. These data provide direct genetic evidence that the point mutations in FGFR3 cause human skeletal dysplasias and uncover a mechanism through which the FGFR3 signals regulate bone growth by modulating expression of Stats and ink4 cell cycle inhibitors.

Amino Acid Substitution↗
PubMed1999

Blood pressure and heart rate in the ovine fetus: ontogenic changes and effects of fetal adrenalectomy.

Ontogenic changes in baseline and 24-h rhythms of fetal arterial blood pressure (FABP) and heart rate (FHR) and their regulation by the fetal adrenal were studied in 18 fetal sheep chronically instrumented at 109-114 days gestation (GA). In the long-term study, FABP and FHR were continuously recorded from 120 days GA to spontaneous term labor (>145 days GA) in five animals. Peak times (PT) and amplitudes (Amp) of cosinor analysis were compared at 120-126, 127-133, and 134-140 days GA. Consistent, significant linear increases in FABP and linear decreases in FHR were observed in all fetuses. Significant 24-h rhythms in FABP and FHR were observed during all the time windows. In the adrenalectomy study, to test the hypothesis that fetal cortisol plays a key role in cardiovascular maturation, fetal adrenals were removed in eight animals (ADX); sham fetal adrenalectomy was performed on five animals (Con). Cortisol (4 microgram/min) was infused intravenously in four ADX fetuses from day 7 postsurgery for 7 days (ADX+F). No significant changes in PT and Amp in FABP and FHR were observed. Plasma cortisol levels remained low in Con and ADX fetuses (<4.9 ng/ml). Cortisol infusion increased fetal plasma cortisol to 22.3 +/- 3.2 ng/ml (mean +/- SE) on day 13 in ADX+F fetuses. FABP increased in control and ADX+F but not ADX fetuses; FHR decreased in control and ADX but rose in ADX+F fetuses. These results suggest that, in chronically instrumented fetal sheep at late gestation, 1) increases in FABP and decreases in FHR are maintained consistently from 120 to 140 days GA, with distinct 24-h rhythms, the PT and Amp of which remain unchanged, and 2) the physiological increase in FABP is dependent on the fetal adrenal; bilateral removal of the fetal adrenals does not prevent the ability of cortisol to produce a sustained increase in FABP.

Adrenal Glands↗
PubMed1999

Early activation of transcription factor NF-kappaB during ischemia in perfused rat heart.

The transcription factor nuclear factor kappaB (NF-kappaB) regulates multiple immediate-early gene expressions involved in immune and inflammatory responses and cellular defenses. Ischemia-reperfusion induces many immediate-early gene expressions, but little is known about the NF-kappaB activation in myocardium during ischemia and reperfusion. This study demonstrated that ischemia alone rapidly induced NF-kappaB activation in the myocardium of isolated working rat hearts. Electrophoretic mobility shift assay showed that NF-kappaB binding activity significantly increased in the nucleus after 5 min of ischemia and remained elevated for up to 30 min. Western blot analysis suggested that the levels of inhibitory IkappaBalpha protein in the cytoplasm became markedly decreased at 4, 5, 7.5, and 10 min of ischemia but were gradually restored following 10 min of ischemia. Reduction of IkappaBalpha protein in the cytoplasm by ischemia resulted in NF-kappaB translocation to the nucleus. Northern blot hybridization showed that IkappaBalpha mRNA levels were not significantly elevated during myocardial ischemia. Pyrrolidine dithiocarbamate, an antioxidant, significantly inhibited the loss of IkappaBalpha protein from the cytoplasm and prevented NF-kappaB binding activity in the nucleus. Reperfusion following short periods of ischemia augmented NF-kappaB binding activity in the nucleus induced by ischemia. The results suggest that early activation of NF-kappaB induced by ischemia in the myocardium could be a signal mechanism for controlling and regulating immediate-early gene expression during ischemia-reperfusion.

Animals↗
PubMed1999

Critical role of glass fiber length in TNF-alpha production and transcription factor activation in macrophages.

Recent studies have demonstrated that dielectrophoresis is an efficient method for the separation of fibers according to fiber length. This method allows the investigation of fiber-cell interactions with fiber samples of the same composition but of different lengths. In the present study, we analyzed the effects of length on the interaction between glass fibers and macrophages by focusing on production of the inflammatory cytokine tumor necrosis factor (TNF)-alpha in a mouse macrophage cell line (RAW 264.7). The underlying molecular mechanisms controlling TNF-alpha production were investigated at the gene transcription level. The results show that glass fibers induced TNF-alpha production in macrophages and that this induction was associated with activation of the gene promoter. Activation of the transcription factor nuclear factor (NF)-kappaB was responsible for this induced promoter activity. The inhibition of both TNF-alpha production and NF-kappaB activation by N-acetyl-L-cysteine, an antioxidant, indicates that generation of oxidants may contribute to the induction of this cytokine and activation of this transcription factor by glass fibers. Long fibers (17 micrometer) were significantly more potent than short fibers (7 micrometer) in inducing NF-kappaB activation, the gene promoter activity, and the production of TNF-alpha. This fiber length-dependent difference in the stimulatory potency correlated with the fact that macrophages were able to completely engulf short glass fibers, whereas phagocytosis of long glass fibers was incomplete. These results suggest that fiber length plays a critical role in the potential pathogenicity of glass fibers.

Acetylcysteine↗
PubMed1999

In vitro antimicrobial activity of a new antiseptic central venous catheter.

A central venous catheter coated with a new antiseptic combination, silver chloride (AgCl) and benzalkonium chloride (BKC) in a polymer matrix, was developed. The antimicrobial efficacy and the ability to prevent surface colonization, after elution in both serum and saline, were evaluated and compared to catheters coated with silver sulfadiazine/chlorhexidine. The results of in vitro assays demonstrated that the AgCl-BKC coated catheters had a broad spectrum of activity against bacteria and C. albicans and prolonged antimicrobial activity for extraction periods of up to 30 days. These data suggest that AgCl-BKC coated catheters may provide another solution for reduction of catheter-related infections.

Anti-Bacterial Agents↗
PubMed1999

Neuropeptide Y and tuberoinfundibular dopamine activities are altered during lactation: role of prolactin.

During lactation the suckling stimulus increases the activity of two populations of neuropeptide Y (NPY) neurons in the hypothalamus, the caudal portion of the arcuate nucleus (ARH) and the dorsomedial hypothalamus (DMH), and suppresses the activity of TIDA neurons in the ARH. In the present study, an acute resuckling model was used to examine the role of suckling-induced hyperprolactinemia in modulating the activity of these systems. Lactating rats were deprived of their eight-pup litters on day 9 postpartum, and 48 h later, the animals served either as nonsuckled controls (0 pups) or were suckled for 24 h. In addition, some of the resuckled animals received two s.c. injections of bromocriptine (0.5 mg/rat x injection), a dopamine D2 agonist, to inhibit suckling-induced PRL secretion. In situ hybridization was performed for rat NPY messenger RNA (mRNA) and tyrosine hydroxylase (TH) mRNA to provide an index for NPY and TIDA neuronal activities, respectively. Resuckling for 24 h induced a significant increase in NPY mRNA levels in the caudal portion of the ARH and in the DMH. Bromocriptine treatment did not alter the increase in NPY mRNA levels in the ARH, whereas the treatment greatly attenuated the increase in NPY mRNA in the DMH. TH mRNA levels in the rostral ARH area returned to basal levels in the nonsuckled control animals, and 24 h of resuckling significantly suppressed TH mRNA expression in this area. Bromocriptine treatment caused a significant increase in TH mRNA levels compared with those in the eight-pup suckled group. Thus, the results from the present study demonstrate that the suckling stimulus activated the two populations of NPY neurons and suppressed TIDA activity. Suckling-induced hyperprolactinemia did not participate in the increase in ARH NPY activity, whereas it played a major stimulatory role in suckling-induced activation of NPY neurons in the DMH and an inhibitory role in suckling-induced suppression of TIDA activity. The increase in TIDA activity after bromocriptine treatment was unexpected and suggests that the role of PRL in the regulation of TIDA activity is significantly altered during lactation.

Animals↗