Biomedical subjects

C Lu

Publications and source records attributed to C Lu.

At least 19 recordsLinked to original sources

PubMed1998

The structure of the beta-propeller domain and C-terminal region of the integrin alphaM subunit. Dependence on beta subunit association and prediction of domains.

The alphaM subunit of integrin Mac-1 contains several distinct regions in its extracellular segment. The N-terminal region has been predicted to fold into a beta-propeller domain composed of seven beta-sheets each about 60 amino acid residues long, with the I-domain inserted between beta-sheets 2 and 3. The structure of the C-terminal region is unknown. We have used monoclonal antibodies (mAbs) as probes to study the dependence of the structure of different regions of the alphaM subunit on association with the beta2 subunit in the alphaM/beta2 heterodimer. All of the mAbs to the I-domain immunoprecipitated the unassociated alphaM precursor and reacted with the alphaM subunit expressed alone on the surface of COS cells. By contrast, four mAbs to the beta-propeller domain did not react with the unassociated alphaM precursor nor with the uncomplexed alphaM subunit expressed on COS cell surface. The four mAbs were mapped to three subregions in three different beta-sheets, making it unlikely that each recognized an interface between the alpha and beta subunits. These results suggest that folding of different beta-propeller subregions is coordinate and is dependent on association with the beta2 subunit. The segment C-terminal to the beta-propeller domain, residues 599-1092, was studied with nine mAbs. A subset of four mAbs that reacted with the alphaM/beta2 complex but not with the unassociated alphaM subunit were mapped to one subregion, residues 718-759, and five other mAbs that recognized both the unassociated and the complexed alphaM subunit were localized to three other subregions, residues 599-679, 820-882, and 943-1047. This suggests that much of the region C-terminal to the beta-propeller domain folds independently of association with the beta2 subunit. Our data provide new insights into how different domains in the integrin alpha and beta subunits may interact.

Amino Acid Sequence↗

PubMed1998

Platelet-derived endothelial cell growth factor/thymidine phosphorylase expression correlated with tumor angiogenesis and macrophage infiltration in colorectal cancer.

To clarify whether platelet-derived endothelial cell growth factor/thymidine phosphorylase (PD-ECGF/TP) expression in both tumor cells and stromal cells has independent or synergistic effects on tumor angiogenesis and progression and to explore a possible regulator for PD-ECGF/TP expression, immunohistochemical staining was conducted on 148 specimens of colorectal cancer. The microvessel count was significantly correlated with the extent of PD-ECGF/TP expression. Macrophage infiltration in tumors with positive TP was significantly higher than in tumors with negative TP (P < 0.001). The Cox model showed that PD-ECGF/TP expression was an independent prognostic factor, although the microvessel count had a stronger value in determining the patient prognosis.

Adenocarcinoma↗

PubMed1998

Salivary concentrations of atrazine reflect free atrazine plasma levels in rats.

The protein binding of atrazine in plasma and its effect on salivary excretion of atrazine was determined in male Sprague-Dawley rats. The degree of protein binding of atrazine was determined at 3 steady-state plasma concentrations, 50, 150, and 250 microg/L, using an ultrafiltration technique. In total, 48 arterial blood samples were collected from 18 rats; 38 of 48 blood samples had their time-matched whole saliva samples. The average protein binding of atrazine ranged from 18% to 37%; however, it was not significantly different across the 3 steady-state plasma concentrations nor among the individual rats. Overall, 26% of atrazine was bound to plasma proteins and not available for transport from blood into saliva. Protein binding of atrazine in plasma was not correlated with total atrazine plasma concentration nor with free atrazine plasma concentration, which indicates that the protein-bound fraction of atrazine is independent of plasma concentration within the range measured in this study (30-400 microg/L). The average saliva/plasma (S/P) concentration ratio of atrazine increased from 0.7 using total atrazine plasma concentration to 0.94 (S/fP) when free atrazine plasma concentrations calculated as 26% of protein binding was used. Salivary concentration was highly correlated with free atrazine plasma concentration. The results suggest that salivary concentration of atrazine not only reflects its total plasma level but accurately measures the portion of atrazine (free atrazine) in plasma, which may be of toxicological significance.

Animals↗

PubMed1998

FtsZ from Escherichia coli, Azotobacter vinelandii, and Thermotoga maritima--quantitation, GTP hydrolysis, and assembly.

We have cloned the ftsZ genes from Thermotoga maritima and Azotobacter vinelandii and expressed the proteins (TmFtsZ and AzFtsZ) in Escherichia coli. We compared these proteins to E. coli FtsZ (EcFtsZ), and found that several remarkable features of their GTPase activities were similar for all three species, implying that these characteristics may be universal among FtsZs. Using a calibrated protein assay, we found that all three FtsZs bound 1 mole guanine nucleotide per mole FtsZ and hydrolyzed GTP at high rates (> 2 GTP per FtsZ per min). All three required magnesium and a monovalent cation for GTP hydrolysis. Previous reports showed that EcFtsZ (and some other species) required potassium. We confirmed this specificity for EcFtsZ but found that potassium and sodium both worked for Az- and TmFtsZ. Specific GTPase activity had a striking dependence on FtsZ concentration: activity (per FtsZ molecule) was absent or low below 50 microg/ml, rose steeply from 50 to 300 microg/ml and plateaued at a constant high value above 300 microg/ml. This finding suggests that the active state requires a polymer that is assembled cooperatively at 50-300 microg/ml. A good candidate for the active polymer was visualized by negative stain electron microscopy--straight protofilaments and protofilament pairs were seen under all conditions with active GTPase. We suggest that the GTP hydrolysis of FtsZ may be coupled to assembly, as it is for tubulin, with hydrolysis occurring shortly after an FtsZ monomer associates onto a protofilament end. As a part of this study, we determined the concentration of EcFtsZ and TmFtsZ by quantitative amino acid analysis and used this to standardize the bicinchonic acid colorimetric assay. This is the first accurate determination of FtsZ concentration. Using this standard and quantitative Western blotting, we determined that the average E. coli cell has 15,000 molecules of FtsZ, at a concentration of 400 microg/ml. This is just above the plateau for full GTPase activity in vitro.

Actin Cytoskeleton↗

PubMed1998

Impact of chronic patency of infarct-related coronary artery on prevalence of myocardial ischemia during the pharmacologic and exercise stress test.

BACKGROUND: Even late restoration of anterograde coronary flow may have beneficial effects on left ventricular function, electrophysiology, and survival in postinfarction patients. HYPOTHESIS: The patency or occlusion of an infarct-related coronary artery in the chronic phase may also be associated with myocardial ischemia provoked by pharmacologic and physiologic stress tests. METHODS: High-dose dipyridamole echocardiography test (DET) (up to 0.84 mg/kg over 10 min), exercise electrocardiography (EET), and coronary angiographic data in a group of 127 in-hospital patients who had survived an acute myocardial infarction were analyzed. Patients who had only angiographic evidence of infarct-related single artery disease (> or = 50% luminal diameter reduction) and no previous revascularization were enrolled in the study. DET and EET were performed (DET in all, EET in 118 patients) within 5 days before coronary angiography. Fifty-seven patients had total occluded infarct arteries (Group 1) with various degrees of collateral circulation (2.6 +/- 1.1 collateral score, by a 3 grading system), whereas the other 70 patients had patent infarct arteries (Group 2) with significant residual stenoses (82 +/- 13% diameter reduction). RESULTS: The prevalence of rest angina or effort angina and topography of the infarct-related coronary artery did not differ between the two groups (all p = NS). There were more patients with Q wave in Group 1 than in Group 2 (72 vs. 57%, p = 0.08) compared with non-Q wave infarction (Group 1 = 28 vs. Group 2 = 43%, p = 0.08). Ischemia in the infarct-related artery territory detected by DET (defined as new wall motion dyssynergy or marked worsening of resting hypokinesia) was 61% in Group 1 and 41% in Group 2 (p = 0.025). EET was positive in 26 of 54 (48%) Group 1 and in 21 of 64 (33%) Group 2 patients (p = 0.09). CONCLUSIONS: Patients with occluded infarct-related arteries have a higher prevalence of ischemia during DET and EET regardless of the presence of collateral flow. These results suggest that the presence of partial anterograde flow in the prolonged period could have a favorable influence on prevalence of residual ischemia in these patients.

Blood Flow Velocity↗

PubMed1998

Maximizing pregnancy rates and limiting higher-order multiple conceptions by determining the optimal number of embryos to transfer based on quality.

OBJECTIVE: To define statistical thresholds for the number of embryos to be transferred to achieve an optimal pregnancy rate and keep higher-order multiple conceptions (pregnancy with more than two fetal sacs with cardiac activity) within an acceptable limit. DESIGN: A retrospective review of patient records. SETTING: Private practice assisted reproductive technology (ART) facility. PATIENT(S): Seven hundred fifty-four consecutive patients who underwent IVF-ET from 1994-1996. INTERVENTION(S): Embryo grading and score system used on day 3 of embryo transfer. MAIN OUTCOME MEASURE(S): Implantation, pregnancy, and multiple conception rates. RESULT(S): For women < or =35 years old, transfer of up to four poor-quality, two fair-quality, or two good-quality embryos is optimal to eliminate any risk of higher-order multiple pregnancies. Transfer of four poor-quality, three fair-quality, or two good-quality embryos is recommended for women 36 to 39 years old. In women who are > or =40 years old, five embryos need to be transferred regardless of embryo quality. CONCLUSION(S): The mean cumulative embryo score can be used as a reference to determine an optimal number of embryos to transfer and to predict pregnancy outcome.

Adult↗

PubMed1998

Actin depolymerization affects stress-induced translational activity of potato tuber tissue

Changes in polymerized actin during stress conditions were correlated with potato (Solanum tuberosum L.) tuber protein synthesis. Fluorescence microscopy and immunoblot analyses indicated that filamentous actin was nearly undetectable in mature, quiescent aerobic tubers. Mechanical wounding of postharvest tubers resulted in a localized increase of polymerized actin, and microfilament bundles were visible in cells of the wounded periderm within 12 h after wounding. During this same period translational activity increased 8-fold. By contrast, low-oxygen stress caused rapid reduction of polymerized actin coincident with acute inhibition of protein synthesis. Treatment of aerobic tubers with cytochalasin D, an agent that disrupts actin filaments, reduced wound-induced protein synthesis in vivo. This effect was not observed when colchicine, an agent that depolymerizes microtubules, was used. Neither of these drugs had a significant effect in vitro on run-off translation of isolated polysomes. However, cytochalasin D did reduce translational competence in vitro of a crude cellular fraction containing both polysomes and cytoskeletal elements. These results demonstrate the dependence of wound-induced protein synthesis on the integrity of microfilaments and suggest that the dynamics of the actin cytoskeleton may affect translational activity during stress conditions.

Journal Article↗

PubMed1997

Functional coupling of a human retinal metabotropic glutamate receptor (hmGluR6) to bovine rod transducin and rat Go in an in vitro reconstitution system.

The cDNA encoding hmGluR6, appended with a 15-amino acid antibody epitope (1D4), was transiently transfected in COS-7 cells. The receptor was purified from COS cell membranes using an antibody affinity column. The purified receptor was then reconstituted into lipid vesicles, and its ability to activate either transducin, the rod photoreceptor-specific GTP-binding protein, or the alpha subunit of Go was assayed in vitro using a guanosine 5'-3-O-(thio)triphosphate binding assay. Activation of both transducin and Go was observed. The rate of Go activation was 18-fold greater than the rate of transducin activation. This indicates that the coupling of mGluR6 to Go is more efficient and suggests that Go may be involved in coupling to mGluR6 in ON-bipolar cells.

Amino Acid Sequence↗

PubMed1997

Tissue-specific regulation of G-protein-coupled inwardly rectifying K+ channel expression by muscarinic receptor activation in ovo.

We investigated the effects of muscarinic acetylcholine receptor stimulation on the expression levels of the G-protein-coupled inwardly rectifying K+ channel (GIRK) subunits using solution hybridization and immunoblot analyses. We report here that treatment of chick embryos in ovo with muscarinic agonist causes decreases in mRNA levels encoding GIRK1 and GIRK4 in atria but does not alter GIRK1 expression in ventricles. In addition, GIRK1 protein levels also demonstrate a decrease in atria upon muscarinic acetylcholine receptor stimulation. Numerous receptors couple to the activation of the GIRK family of inwardly rectifying K+ channels; thus, these decreases represent a novel mechanism for regulating physiological responses to chronic agonist exposure.

Animals↗

PubMed1997

Association of vascular endothelial growth factor expression with tumor angiogenesis, survival and thymidine phosphorylase/platelet-derived endothelial cell growth factor expression in human colorectal cancer.

To examine the association of vascular endothelial growth factor (VEGF) expression with tumor angiogenesis, survival and thymidine phosphorylase/platelet-derived endothelial cell growth factor (dThdPase/PD-ECGF) expression in human colorectal cancer, immunohistochemical studies were performed on 136 cases of resected colorectal cancer specimens using antibodies for VEGF, KDR, CD34 and dThdPase/PD-ECGF. Fifty-nine cases (43%) were evaluated as positive for VEGF staining and 71 cases (52%) were evaluated as positive for dThdPase/PD-ECGF staining. The expression of VEGF correlated significantly with vessel counts and the expression of dThdPase/PD-ECGF (P = 0.01 and 0.01, respectively). Cox proportional hazards model analysis showed that vessel counts and VEGF expression were significant and independent prognostic factors, but that KDR expression was not.

Adult↗

PubMed1997

Stimulus-secretion coupling in porcine adrenal chromaffin cells: effect of dexamethasone.

Recent studies from this laboratory established that dexamethasone (DEX) potentiates Ca2+ current via voltage-gated Ca2+ channels (VGCC), and as a consequence potentiates agonist-induced cytosolic Ca2+ transients in rat adrenal chromaffin cells. The present study examined whether DEX can also modulate VGCC activity and agonist-induced cytosolic Ca2+ transients in porcine adrenal medullary chromaffin (PAMC) cells, and if so whether this results in alterations in catecholamine secretion. Forty-eight-hr exposure to 1 microM DEX significantly increased peak Ca2+ current (delta + 138%; n = 6; P < 0.05) in PAMC cells. DEX treatment also significantly potentiated the increase in cytosolic Ca2+ in response to membrane depolarization with KCl (delta + 20%; n = 29; P < 0.05), but did not affect the amplitude of Ca2+ transients elicited by nicotine or acetylcholine. Despite the potentiation of intracellular Ca2+, DEX treatment had no effect on KCl-induced secretion of either norepinephrine or epinephrine. These data demonstrate that as in the rat chromaffin cell, DEX can also increase VGCC activity in PAMC cells. However, the subsequent potentiation of selected agonist-induced increases in intracellular Ca2+ does not appear to be sufficient to alter catecholamine secretion.

Acetylcholine↗

PubMed1997

Channel activators regulate ATP-sensitive potassium channel (KIR6.1) expression in chick cardiomyocytes.

ATP-sensitive potassium channels (K(ATP)) are widely expressed and yet little is known about the mechanisms regulating their expression. Here we report that expression of chick heart Kir6.1 is regulated by channel activators. Activation of K(ATP) with either ATP depletion or pinacidil, up-regulated Kir6.1 mRNA 1.8- to 2.4-fold in cultured ventricular myocytes as measured by competitive PCR. Pinacidil treatment also increased Kir6.1 protein as detected using an antibody to Kir6.1. Glibenclamide, a K(ATP) inhibitor, completely blocked the pinacidil-induced increase in Kir6.1 levels. It appears that Kir6.1 is up-regulated by an unknown signal transduction pathway initiated by K(ATP) opening.

Adenosine Triphosphate↗

PubMed1997

Folding of the conserved domain but not of flanking regions in the integrin beta2 subunit requires association with the alpha subunit.

We have used immunoprecipitation with mAbs to probe folding during biosynthesis of the beta2 integrin subunit of lymphocyte function-associated antigen 1 (LFA-1; CD11a/CD18) before and after association with the alphaL subunit. An evolutionarily conserved region is present in the beta2 subunit between amino acid residues 102 and 344. mAbs to one subregion before the conserved region, and two subregions after the conserved domain, immunoprecipitated both the unassociated beta'2 precursor and mature alphaL/beta2 complex, suggesting portions of these subregions are folded before association with alphaL. An activating mAb to the C-terminal cysteine-rich region, KIM127, preferentially bound to the unassociated beta subunit, suggesting that it may bind to an epitope that is in an alphabeta interface in unactivated LFA-1. By contrast, mAbs to five different epitopes in the conserved region did not react with unassociated beta'2 precursor, suggesting that this region folds after alphaL association and is intimately associated with the alphaL subunit in the alphaL/beta2 complex. mAbs to two different epitopes that involve the border between the conserved region and the C-terminal segment, were fully or partially reactive with the beta'2 precursor, suggesting that this region is partially folded before association with alphaL. The findings suggest that the conserved region is a distinct folding and hence structural unit, and is intimately associated with the alpha subunit.

Amino Acid Sequence↗

PubMed1997

Effect of dexamethasone on voltage-gated Ca2+ channels and cytosolic Ca2+ in rat chromaffin cells.

The present study examined whether the synthetic glucocorticoid dexamethasone (DEX) can modulate voltage-gated Ca2+ channel (VGCC) activity, and as a consequence agonist-induced increases in cytosolic Ca2+, in cultured rat adrenal medullary chromaffin (RAMC) cells. Exposure to 1 microM DEX for 48 h significantly increased peak VGCC current (delta +140%). DEX treatment also significantly potentiated the increases in cytosolic Ca2+ in response to submaximal stimulatory concentrations of KCl (delta +64%) and nicotine (delta +32%). The Ca2+ channel agonist BAY K-8644 increased both VGCC current (delta +109%) and potentiated the KCl-stimulated increase in cytosolic Ca2+ (delta +35%) to a comparable extent to that seen with DEX. These data suggest that DEX treatment increases VGCC activity, and that this increased Ca2+ influx leads to potentiation of agonist-induced increases in cytosolic Ca2+ in RAMC cells.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗

PubMed1997

Modulation of hybrid bass retinal gap junctional channel gating by nitric oxide.

1. To elucidate the role of the nitric oxide (NO) transmitter system in the regulation of gap junctional channel gating, we have examined the effects of the NO donor sodium nitroprusside (SNP) on the electrical synapses of hybrid bass H2-type horizontal cells. 2. SNP reversibly reduced the macroscopic junctional conductance without significantly changing voltage sensitivity. 3. Kinetic analyses showed that SNP made the voltage-dependent decay of junctional currents more rapid. 4. Single-channel data showed that SNP reduced channel open probability by reducing channel open frequency. 5. The action of SNP can be prevented or largely reduced by the NO scavenger, haemoglobin. NO release by SNP solutions was detected directly by a NO sensor. 6. NO appears to modulate the gap junctional conductance by activating the cGMP-cGMP-dependent protein kinase G (PKG) pathway. A membrane-permeable cGMP analogue, 8-Br-cGMP, mimics the action of SNP. A soluble guanylate cyclase inhibitor (LY-83583) and a highly specific cGMP-dependent protein kinase inhibitor (RKRARKE) blocked the action of NO. 3-Isobutyl-1-methylxanthine (IBMX), a non-specific phosphodiesterase inhibitor, potentiated the effect of SNP. 7. [Ca2+]i image studies showed that NO donors did not change [Ca2+]i in horizontal cells, suggesting that the regulation of junctional channels by NO is [Ca2+]i independent.

1-Methyl-3-isobutylxanthine↗

PubMed1997

Tumor angiogenesis and mode of metastasis in patients with colorectal cancer.

Neovascularization promotes tumor growth by facilitating nutrient exchange and by the paracrine effect. To investigate the relationship between tumor angiogenesis and patient outcome in colorectal cancer, 133 primary tumors were immunostained for CD34 antigen. Blood vessels within five microscopic fields at x200 were counted, and the mean was assigned. Mean patient age was 62.9 years, mean follow-up was 56.4 months, and mean vessel count was 112 (range, 23-298). Cox proportional hazards model and multivariate logistic regression analyses showed that the vessel count was the most important prognostic factor and correlated significantly with hematogenous, but not peritoneal or lymph node, metastasis.

Adenocarcinoma↗

PubMed1997

Determination of atrazine levels in whole saliva and plasma in rats: potential of salivary monitoring for occupational exposure.

Current biological monitoring techniques are often unable to provide accurate estimates of pesticide dose in exposed worker populations. This study was conducted to investigate the feasibility of pesticide biomonitoring using saliva. Atrazine [2-chloro-4-ethylamino-6-(isopropylamino)-s-triazine], a member of the triazine herbicides, was selected to investigate salivary excretion following direct gastric administration in rats. Concentrations of atrazine in whole saliva and arterial plasma samples were determined by enzyme-linked immunosorbent assay (ELISA). Atrazine reached its highest level in both arterial plasma (238 micrograms/L) and whole saliva (157 micrograms/L) 35 min after administration of 105 mg/kg of atrazine, and then decreased with time in a parallel fashion. Although saliva atrazine levels were lower than levels in arterial plasma, there was a very high correlation between whole saliva and arterial plasma atrazine concentrations (r2 = .95). In addition, pharmacokinetic analysis suggested that salivary levels of atrazine can be used to predict concentrations of atrazine in plasma. The mean whole saliva/arterial plasma atrazine concentration ratio (S/P) was 0.66 +/- 0.11 (n = 20). The S/P ratios did not vary significantly over time, and were not affected by salivary flow rate. This study demonstrates that atrazine is transported into saliva, and that a relatively constant concentration ratio between whole saliva and arterial plasma is maintained. Because the salivary concentrations of atrazine are independent of variation in salivary flow rate, salivary monitoring of atrazine in humans may prove useful and practical. Finally, this study suggests that other pesticides with chemical and physical properties similar to those of atrazine can be monitored in saliva.

Animals↗

PubMed1997

Spontaneous apoptosis is inversely related to intratumoral microvessel density in gastric carcinoma.

Increasing evidence from experimental studies indicates that apoptosis may be related to angiogenesis in tumor progression. To explore how spontaneous apoptosis correlates with tumor angiogenesis, we measured the apoptotic index (AI) using the ApopTag kit (Oncor) and intratumoral microvessel density using an anti-CD34 monoclonal antibody in 101 cases of gastric carcinoma. Immunohistochemical staining for Ki-67 labeling index and the expression of p53 were conducted simultaneously. Statistical analysis revealed an inverse correlation between AIs and intratumoral microvessel densities (r = -0.4066, P < 0.0001) and failed to find significant correlations between AI and Ki-67 labeling index, as well as the expression of p53. The results of this study demonstrated for the first time that the incidence of apoptosis in gastric carcinoma is significantly influenced by the extent of neovascularization and suggests that tumor angiogenesis may contribute to a reduction of apoptosis in tumor cells.

Apoptosis↗