Biomedical subjects

S Lin

Publications and source records attributed to S Lin.

At least 37 records · Page 2Linked to original sources

PubMed1996

Structural requirements for targeting of surfactant protein B (SP-B) to secretory granules in vitro and in vivo.

Human surfactant protein B (SP-B) is synthesized by type II cells as a 381-residue preproprotein which is proteolytically processed to a 79-residue mature peptide and targeted to lamellar bodies for secretion. To identify secretory granule targeting determinants, constructs encoding the SP-B preproprotein (SP-B), COOH-terminally deleted SP-B (SP-BDeltaC), the NH2-terminal propeptide (SP-BN), and a chimeric molecule consisting of albumin and the mature peptide (ALB/SP-BM) were transfected into AtT-20 and PC12 cells. Pulse-chase studies demonstrated that 10-30% of SP-B and SP-BDeltaC remained in cells in an endoglycosidase H-resistant form. Secretion of stored SP-B was stimulated by forskolin/12-O-tetradecanoylphorbol-13-acetate and intracellular SP-B was localized to secretory granules by immunoelectron microscopy. In contrast, SP-BN and ALB/SP-BM were constitutively secreted and not detected in secretory granules. Specific processing of SP-B was not detected in either AtT-20 or PC12 cells. Expression of SP-BDeltaC in transgenic mice resulted in secretion of fully processed mature SP-B, indicating correct processing and targeting of this construct in vivo. We conclude that 1) SP-B processing occurs in a cell-specific manner, 2) the proprotein contains secretory granule targeting determinants that are not cell-specific, 3) the NH2-terminal propeptide and the mature peptide are required for targeting SP-B to lamellar body, and 4) the COOH-terminal propeptide is not required for processing or sorting of SP-B.

Animals↗

PubMed1996

Novel azacyclic ureas that are potent inhibitors of HIV-1 protease.

A series of novel, azacyclic ureas which are highly potent inhibitors of the HIV-1 protease (IC50 = 4.1 to < 0.5 nM) were synthesized. Aqueous solubilities of this series of compounds were improved by incorporating polar functional groups at the P1' P2 and P2' positions. These compounds also possess good anti-viral activity by inhibition of the cytopathic effect of HIV-13B in MT-4 cells in vitro.

Amino Acid Sequence↗

PubMed1996

Interdigitation of bilayers from ether lipid analogs: (R)-PAF, (R)-Lyso-PAF and the antineoplastic (R)-ET-18-OMe.

The ether lipid analogs (R)-PAF, (R)-ET-18-OMe and (R)-Lyso-PAF can form bilayers. Using differential scanning calorimetry (DSC) and small angle X-ray diffraction, we have investigated bilayers formed entirely of these molecules. Small angle X-ray diffraction experiments strongly indicated that these bilayers are interdigitated at 98% relative humidity in the gel phase. The slight differences in the structures of the ether lipids affect their thermal behavior and packing properties. Of the three, (R)-Lyso-PAF shows the highest phase transition temperature, broadest phase transition peak and smallest enthalpy change. These characteristics may be attributed to hydrogen bonding of the -OH group at sn-2 position of its glycerol backbone to water or to other (R)-Lyso-PAF molecules. The interdigitated structure shows that these bilayers are similar to lipid bilayers and suggests that these molecules could easily insert themselves into membranes to reach their respective active sites or to perturb the membranes. The packing and thermal characteristics of (R)-Lyso-PAF may help explain its biological inactivity.

Antineoplastic Agents↗

PubMed1996

Characterization of the structure and function of a new mitogen-activated protein kinase (p38beta).

Mitogen-activated protein (MAP) kinase cascades represent one of the major signal systems used by eukaryotic cells to transduce extracellular signals into cellular responses. Four MAP kinase subgroups have been identified in humans: ERK, JNK (SAPK), ERK5 (BMK), and p38. Here we characterize a new MAP kinase, p38beta. p38beta is a 372-amino acid protein most closely related to p38. It contains a TGY dual phosphorylation site, which is required for its kinase activity. Like p38, p38beta is activated by proinflammatory cytokines and environmental stress. A comparison of events associated with the activation of p38beta and p38 revealed differences, most notably in the preferred activation of p38beta by MAP kinase kinase 6 (MKK6), whereas p38 was activated nearly equally by MKK3, MKK4, and MKK6. Moreover, in vitro and in vivo experiments showed a strong substrate preference by p38beta for activating transcription factor 2 (ATF2). Enhancement of ATF2-dependent gene expression by p38beta was approximately20-fold greater than that of p38 and other MAP kinases tested. The data reported here suggest that while closely related, p38beta and p38 may be regulated by differing mechanisms and may exert their actions on separate downstream targets.

Activating Transcription Factor 2↗

PubMed1996

Structural requirements for intracellular transport of pulmonary surfactant protein B (SP-B).

Human SP-B is synthesized by the alveolar Type II epithelial cell as a 381 amino acid preproprotein. The 79 residue mature SP-B peptide is extremely hydrophobic and flanked by propeptides of 200 and 102 amino acids at its NH2- and COOH-termini, respectively. The purpose of this study was to identify peptide domains of the SP-B proprotein necessary for trafficking of the mature peptide in the secretory pathway. To this end several constructs were generated, by subcloning the full length human SP-B (SP-B), COOH-terminally truncated SP-B (SP-B delta C, in which residues 201-381 were deleted), NH2-terminally deleted SP-B (SP-B delta N, in which residues 28-200 were deleted), NH2-terminal propeptide (SP-BN), mature SP-B (SP-BM) and COOH-terminal propeptide (SP-BC), into the mammalian expression vector pcDNA3. The resulting expression constructs were characterized by DNA sequencing and in vitro transcription/translation and subsequently transfected into Chinese hamster ovary cells. 48 h after transfection, cells were labeled with [35S]-met/cys and analyzed by immunoprecipitation, SDS-PAGE and autoradiography. Proteins encoded by SP-B, SP-B delta C, SP-BN and SP-BC constructs were secreted into media; in contrast, SP-B constructs lacking the NH2-terminal propeptide (SP-B delta N) remained in the endoplasmic reticulum (as assessed by endoglycosidase H sensitivity) and were rapidly degraded. We conclude that (1) 27 amino acids at the NH2-terminus of SP-B contain a functional signal peptide and (2) the NH2-terminal propeptide of the SP-B precursor is necessary and sufficient for intracellular trafficking of the mature peptide.

Animals↗

PubMed1996

The Fc(epsilon)RIbeta subunit functions as an amplifier of Fc(epsilon)RIgamma-mediated cell activation signals.

The high affinity IgE receptor (Fc(epsilon)RI) plays a central role in the initiation of allergic responses. Fc(epsilon)RI is multimeric and is expressed as either (alpha)(gamma2) trimers or (alpha)(beta)(gamma2) tetramers. Recently, polymorphisms of the beta chain gene have been associated with the development of various allergic phenotypes. Until now, the role of beta in Fc(epsilon)RI-dependent signaling was largely unknown. For that reason, we compared the signaling characteristics of various wild-type and mutant (alpha)(gamma2) and (alpha)(beta)(gamma2) Fc(epsilon)RI complexes. These studies demonstrate that the gamma dimer functions as an autonomous activation module, while beta functions as an amplifier that provides a gain of 5- to 7-fold, as measured by Syk activation and calcium mobilization.

Calcium↗

PubMed1996

High-performance liquid chromatographic determination of anandamide amidase activity in rat brain microsomes.

A rapid, sensitive, and reliable method for measuring anandamide amidase activity in rat brain microsomes by reversed-phase high-performance liquid chromatography (RP-HPLC) and its applications are described. Enzymatic activity was assayed by the determination of the rates of hydrolysis of anandamide or its analogs at 37 degrees C. The reaction products were separated using an ODS guard column eluted with aqueous phosphoric acid-acetonitrile and quantitated with uv detection at 204 nm and an external standard method. Baseline separation of the acid products from their substrates was completed in less than 2 min. The detection limits were 1.4 pmol for arachidonic acid and 0.22 pmol for anandamide at a signal to noise ratio of 4:1. The stability of anandamide in the acidic mobile phase was tested, and no significant decomposition was observed up to 1 h. The method was successfully applied to the examination of substrate specificity as well as for testing the ability of amidase inhibitors to block its hydrolysis. Kinetic constants obtained for (S)-methanandamide were an apparent Km of 8.6 +/- 1.3 microM and a Vmax of 362 +/- 16 pmol/min/mg of protein. A highly potent inhibitor, palmitylsulfonyl fluoride (PSF), was found to have an IC50 of 50 nM. PSF is 210 times as potent as phenylmethylsulfonyl fluoride. The method offers several advantages over existing methodology using radioisotopes or a solvent extraction procedure.

Amidohydrolases↗

PubMed1996

Low-temperature femtosecond-resolution transient absorption spectroscopy of large-scale symmetry mutants of bacterial reaction centers.

Reaction centers isolated from three large-scale symmetry mutants sym0, sym2-1, and sym5-2 described in the previous article of this issue [Taguchi, A. K. W., Eastman, J. E., Gallo, D. M., Jr., Sheagley, E.. Xiao, W., & Woodbury, N. W. (1996) Biochemistry 35, 3175-3186] have been investigated by low-temperature ground state and ferntosecond-resolution transient absorption spectroscopy. All three of these large-scale symmetry mutants undergo electron transfer at 20 K. The mutants sym0 and sym5-2 have yields and dominant rates of charge separation comparable to wild type. However. the sym2-mutant shows a roughly 35%, quantum yield at this temperature, and the major kinetic component of the initial electron transfer is slower than wild type by nearly a factor of 100. The sym0 mutant showed substantial changes in the monomer bacteriochiorophyll ground state and transient spectra, and both sym0 sym2-1 showed changes in the bacteriopheophyll ground state and transient spectra. In particular, sym2-1 shows a small absorbance decrease in the region of the Qx band of the B side bacteriopheophytin which could be attributed to 10%-20% electron transfer along the B pathway.

Cold Temperature↗

PubMed1996

Activation of BTK by a phosphorylation mechanism initiated by SRC family kinases.

Bruton's tyrosine kinase (BTK) is pivotal in B cell activation and development through its participation in the signaling pathways of multiple hematopoietic receptors. The mechanisms controlling BTK activation were studied here by examination of the biochemical consequences of an interaction between BTK and SRC family kinases. This interaction of BTK with SRC kinases transphosphorylated BTK on tyrosine at residue 551, which led to BTK activation. BTK then autophosphorylated at a second site. The same two sites were phosphorylated upon B cell antigen receptor cross-linking. The activated BTK was predominantly membrane-associated, which suggests that BTK integrates distinct receptor signals resulting in SRC kinase activation and BTK membrane targeting.

3T3 Cells↗

PubMed1996

A novel, picomolar inhibitor of human immunodeficiency virus type 1 protease.

The design, synthesis, and molecular modeling studies of a novel series of azacyclic ureas, which are inhibitors of human immunodeficiency virus type 1 (HIV-1) protease that incorporate different ligands for the S1', S2, and S2' substrate-binding sites of HIV-1 protease are described. The synthesis of this series is highly flexible in the sense that the P1', P2, and P2' residues of the inhibitors can be changed independently. Molecular modeling studies on the phenyl ring of the P2 and P2' ligand suggested incorporation of hydrogen-bonding donor/acceptor groups at the 3' and 4-positions of the phenyl ring should increase binding potency. This led to the discovery of compound 7f (A-98881), which possesses high potency in the HIV-1 protease inhibition assay and the in vitro MT-4 cell culture assay (Ki = approximately 5 pM and EC50 = 0.002 microM). This compares well with the symmetrical cyclic urea 1 pioneered at DuPont Merck.

Binding Sites↗

PubMed1996

Schizophrenia after prenatal famine. Further evidence.

BACKGROUND: Suggestive findings of an earlier study that prenatal nutritional deficiency was a determinant of schizophrenia prompted us to undertake a second test of the hypothesis using more precise data on both exposure and outcome. METHODS: Among persons born in the cities of western Netherlands during 1944 through 1946, we compared the risk for schizophrenia in those exposed and unexposed during early gestation to the Dutch Hunger Winter of 1944/1945. The frequency of hospitalized patients with schizophrenia at age 24 to 48 years in the exposed and unexposed birth cohorts was ascertained from a national psychiatric registry. RESULTS: The most exposed birth cohort, conceived at the height of the famine, showed a twofold and statistically significant increase in the risk for schizophrenia (relative risk [RR] = 2.0; 95% confidence interval [CI] = 1.2 to 3.4; P < .01) in both men (RR = 1.9; 95% CI = 1.0 to 3.7; P = .05) and women (RR = 2.2; 95% CI = 1.0 to 4.7; P = .04). Among all birth cohorts of 1944 through 1946, the risk for schizophrenia clearly peaked in this exposed cohort. CONCLUSION: Prenatal nutritional deficiency may play a role in the origin of some cases of schizophrenia.

Adult↗

PubMed1996

Immunoquantitation and microsomal monooxygenase activities of hepatic cytochromes P4501A and P4502B and chlorinated hydrocarbon contaminant levels in polar bear (Ursus maritimus).

Contamination of the Arctic ecosystem by anthropogenic compounds has resulted in exposure of polar bear (Ursus maritimus) to lipophilic chlorinated hydrocarbon contaminants (CHCs) accumulated through the marine food web. Liver samples were collected from 16 adult male polar bears in the Canadian arctic and subjected to chemical analysis for CHCs and metabolites, determination of alkoxyresorufin O-dealkylase activities, and immunoquantitation of cytochrome P450 (CYP) protein levels. We report on the relationships between the hepatic microsomal levels of immunoreactive CYP1A and CYP2B isozymes, catalytic activities, and hepatic CHC and metabolite concentrations in polar bear. We specifically explored the influence of several CHCs on the induction of hepatic CYP in polar bear and the potential use of immunoassay quantitation as a bioindicator of CHC exposure. Polychlorinated biphenyls (PCB) classed as CYP1A and mixed CYP1A/CYP2B inducers accounted for about 25% of the total PCB residues present (18,680 +/- 5053 ng/g lipid). CYP1A protein content correlated strongly with hepatic levels of PCBs, PCDDs (0.032 +/- 0.018 ng/g lipid, and PCDFs (0.011 +/- 0.007 ng/g lipid) and their corresponding toxic equivalents (TEQ, 0.377 +/- 0.182 ng/g lipid). Mono-ortho-CB-156, CB-157, and CB-105 were the predominant TEQ contributors. Correlations between CYP2B protein content and CHC residue levels in polar bear liver suggested that ortho-chlorine-substituted PCBs and chlordanes were the major contributors to CYP2B induction. CYP1A and CYP2B contents were therefore good indicators of CHC exposure in polar bear liver. Ethoxyresorufin, pentoxyresorufin, and benzyloxyresorufin O-dealkylase activities increased with increasing CYP1A protein content up to protein levels of approximately 5 pmol/mg, suggesting that all three activities were primarily CYP1A-mediated. These results were substantiated by antibody inhibition experiments. In summary, immunoquantitated CYP1A and CYP2B isozymes are a more reliable measure of exposure to CHC inducers than alkoxyresorufin O-dealkylase activities in polar bear.

Animals↗

PubMed1996

Pregnancy outcome following genetic amniocentesis at 11-14 versus 16-19 weeks' gestation.

OBJECTIVE: To compare pregnancy complications in women having genetic amniocentesis at 11-14 weeks versus those undergoing amniocentesis at 16-19 weeks' gestation. METHODS: A genetics data base was used to identify patients retrospectively, those who had genetic amniocenteses by three experienced operators during a 4-year period. The study group consisted of women who had amniocenteses at 11-14 weeks' gestation. For each study patient (early amniocentesis), two controls (amniocentesis at 16-19 weeks) were identified and matched for maternal age, race, and the number of prior spontaneous abortions. An immediate post-procedure complication was defined as any vaginal bleeding, rupture of membranes, or fetal loss occurring up to 30 days after the amniocentesis. A later complication was defined as any fetal death longer than 30 days after the amniocentesis, any preterm delivery, any infant weighing less than the tenth percentile for gestational age, and any neonatal death. Immediate and later complications were compared between the study and control groups. RESULTS. The study group consisted of 314 patients who were matched to 628 controls. Women who had a genetic amniocentesis performed at 11-14 weeks were significantly more likely to have post-procedure amniotic fluid leakage (2.9 versus 0.2%), post-procedure vaginal bleeding (1.9 versus 0.2%), and a fetal loss within 30 days of the amniocentesis (2.2 versus 0.2%) than women undergoing genetic amniocentesis at 16-19 weeks' gestation. Four of the seven patients (57%) with a fetal loss within 30 days of an early amniocentesis had procedure-related complications, such as amniotic fluid leakage, bleeding, and infection, that caused the pregnancy to be lost. No differences were noted between the two groups in the number of preterm deliveries, later fetal deaths, neonatal deaths, or newborns weighing less than the tenth percentile for gestational age. CONCLUSION: Genetic amniocentesis at 11-14 weeks is associated with more post-procedure complications and a higher fetal loss rate within 30 days of the procedure than a genetic amniocentesis performed at 16-19 weeks' gestation.

Adult↗

PubMed1996

Changes in substance P in the jejuna of rats after burns.

Radioimmunoassay (RIA) was used to determine the dynamic changes of immunoreactive substance P (iSP) in the jejuna of rats (TBSA 30 per cent full skin thickness burn) during the first 72 h postburn. Immunohistochemistry and image analysis techniques were used to observe and quantitate the SP immunoreactivity (SP-IR) of positive nerve fibres in the villi of jejuna postburn. Changes in the amount of iSP in jejuna of burned rats were: (1) iSP increased significantly at 1 h postburn, and the high level of iSP lasted 4 h; it then decreased greatly 8 h later with the low level of iSP persisting for 72 h. (2) Significant changes in SP-IR-positive nerve fibres in the villi after burn were shown by the immunohistochemical studies including the morphoses; the distributive densities and SP-IR-positive products in the SP-IR-positive nerve fibres. The results quantified by image analysis showed similar alterations in the distributive densities and SP-IR-positive products in the nerve fibres in the villi during 72 h postburn; which were distinctly elevated by 1 hr then dropped by 8 h and 12 h and finally elevated again. The results indicated that the irritation-release and consumption of SP occurred in jejuna of rats after burns. It might be that SP contributed to the postburn intestinal lesion in rats by bioactivities, such as enhancing the vascular permeability and regulating the intestinal movement. The SP peptidergic nerve fibres of the villus had a direct effect on the damage to mucosal epithelia.

Animals↗

PubMed1996

Requirements for green fluorescent protein detection in transgenic zebrafish embryos.

We have generated transgenic (Tg) lines of zebrafish in which the green fluorescent protein (GFP)-encoding gfp cDNA is driven by the Xenopus laevis ef1 alpha enhancer/promoter; Tg embryos from most of these lines show detectable fluorescence throughout their body. We have investigated the copy number of the Tg genes in fluorescent and non-fluorescent lines, in order to determine how this affects the production of detectable levels of GFP in the zebrafish embryo. Additionally, we have injected purified recombinant GFP into embryos to determine the intracellular GFP concentration required for detection, both when all of the cells in the embryo contain GFP and when only a few do.

Animals↗

PubMed1996

Fertility rates among lead workers and professional bus drivers: a comparative study.

We examined the relationship between lead exposure and fertility among men in a retrospective cohort study. Fertility (1981-1992) of lead-exposed workers was determined from birth certificate information and was compared with that of nonexposed workers. The exposed group consisted of 4256 reproductive-age male workers reported to the New York State Heavy Metals Registry. The comparison group consisted of a random sample of male bus drivers licensed in the state of New York; these men were frequency-matched by age and residence to the men who were exposed to lead. The actual number of births among lead workers was lower than the expected number of births for that group (standardized fertility ratio [SFR] = 0.88; 95% confidence interval [CI] = 0.81-0.95), especially among those who had elevated blood lead levels for longer than 5 years (SFR = 0.43; 95% confidence interval [CI]: = 0.31, 0.59). Even after adjusting for age, race, education, and residence, workers with > 5 years of exposure had reduced likelihood of fathering a child than those with a shorter period of exposure (relative risk, 0.38; 95% CI: 0.23, 0.61). Our study indicates that men with a long duration of lead exposure might have reduced fertility.

Adult↗

PubMed1996

Esophageal bolus transit imaged by ultrafast computerized tomography.

BACKGROUND & AIMS: Conventional transit imaging techniques detect only the solid or liquid component of a swallowed bolus within the esophagus. This study aimed to dynamically image the composition, distribution, and propulsion of esophageal contents during swallowing. METHODS: Multiple adjacent cross-sectional images of the esophagus were obtained in 7 subjects using ultrafast computerized tomography. Images from two 10-mL swallows were synchronized and analyzed for bolus distribution and propulsion, cross-sectional area, intraluminal volume, and intraluminal content. RESULTS: Both liquid and air were swallowed, with the relative distribution varying among levels. Within the ampulla, air occupied 71% of the luminal cross-sectional area. Air was propelled ahead of the liquid bolus at 17 cm/s compared with 7 cm/s for fluid (P < 0.01) and accumulation within the ampulla. Mean bolus velocity was slower through the ampulla. A variable (8-32 mL) volume of air was ingested during swallowing. CONCLUSIONS: Ultrafast computerized tomography studies during transit of a swallowed bolus through the esophagus showed substantial aerophagia and partial bolus separation with air preceding fluid. The ampulla exhibited greater distention that the tubular esophagus because the bolus accumulated at this level before transfer across the hiatus.

Adult↗

PubMed1996

Oropharyngeal accommodation to swallow volume.

BACKGROUND & AIMS: The oropharyngeal swallow accommodates a range of bolus volumes with substantial impact on its dynamic radiographic appearance despite being a nearly reflexive sequence. The aim of this study was to analyze the mechanism of volume accommodation. METHODS: Coordination of the glossopalatal junction, velopharyngeal junction, laryngeal vestibule, and upper esophageal sphincter (UES) and intraluminal pharyngeal dimensions were measured from biplane videofluoroscopic swallowing studies in 8 volunteers during 1- and 20-mL swallows. These measurements were applied to three-dimensional reconstructions of the pharyngeal swallow, permitting analysis of their effect on intraluminal volume and propulsive function. RESULTS: Dividing the pharyngeal swallow into periods of reconfiguration, sustained reconfiguration, and offset of reconfiguration, volume accommodation occurred by a 0.2-second prolongation of reconfiguration, preserving the coordination of the onset and offset events. Augmented reconfiguration increased the dimensions of the pharyngeal chamber, caused more rapid bolus expulsion, and increased UES distention. However, maximal transphincteric flow per unit area across the UES was constant, showing its compliance. CONCLUSIONS: Volume accommodation is accomplished by augmenting and prolonging pharyngeal reconfiguration from a respiratory to deglutitive pathway. This modification in the timing of neurally mediated events is amplified by its mechanical consequences, resulting in the different cineradiographic appearance and propulsive attributes of large and small volume swallows.

Adult↗