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Biomedical subjects

W Lin

Publications and source records attributed to W Lin.

At least 37 records · Page 2Linked to original sources
PubMed2001

Ion pairing between Cl- or ClO4- and alkali metal complexes of ionophore antibiotics in organic solvents: a multinuclear NMR and FT-IR study.

The extent of ion pairing in chloride and perchlorate salts was studied by measurement of the Cl- and ClO4- resonances and the observation of the perchlorate stretching frequency by use of nuclear magnetic resonance (NMR) spectroscopy and Fourier transform infrared spectroscopy (FT-IR), respectively, for a variety of ionophores in various solutions and in large unilaminar vesicles (LUVs). The NMR line widths of chloride and perchlorate were larger in solutions containing the neutral ionophores valinomycin (Val) and nonactin (Non) than in solutions containing the negatively charged ionophores nigericin (Nig), lasalocid (Las), and monensin (Mon). The viscosity-corrected perchlorate NMR line widths in solutions containing Val and Las were significantly negatively correlated (r2 > or = 0.99) with the dielectric constant of the solvent. Solvents with low dielectric constants favored ion pair formation. From methanolic solutions containing the Li+, Na+, K+, and Cs+ salts of Cl- and ClO4-, it was determined that the cation with the highest selectivity for the ionophore affords the most ion pairing. A decrease in pH from 7 to 3 had no significant effect on the NMR line widths of chloride and perchlorate in methanolic solutions containing Val, whereas a similar decrease in pH in a methanolic solution containing Mon caused a 2-fold increase in the line widths. The FT-IR difference spectrum of KClO4 in a methanolic solution containing Val showed splitting at the perchlorate stretching frequency. No band splitting was observed in the FT-IR difference spectrum of KClO(4) in methanolic solutions containing Las. The efflux of 35Cl in LUVs containing the neutral ionophore Val followed first-order kinetics with an efflux constant of 1.70 x 10(-3) x min(-1), as determined by 35Cl NMR spectroscopy. The induction of increased membrane permeability in LUVs by the ionophore was determined to be negligible for Val and Nig by fluorescence spectroscopy.

Anions↗
PubMed2001

Preparation and characterisation of rose Bengal-loaded surface-modified albumin nanoparticles.

Surface-modified albumin nanoparticles were prepared from two poly(ethylene glycol)-human serum albumin conjugates: poly(thioetheramido acid)-poly(ethylene glycol) copolymer-grafted HSA (HSA-PTAAC-PEG) and methoxy poly(ethylene glycol)-grafted HSA (HSA-mPEG). Rose bengal (RB) was used as a model drug for encapsulation into the nanoparticles either during the particle production or by adsorption post particle preparation. The drug incorporation and release was affected by the different production methods and the different polymer compositions. When RB was loaded in HSA and HSA/HSA-PTAAC-PEG nanoparticles, up to 5% (w/w) drug content was achieved. The drug loading in HSA-mPEG nanoparticles was much lower and the results from the microcalorimetry study indicated that the low loading efficiency was due to less drug-protein binding sites available in the HSA-mPEG molecule as compared to the HSA molecule. The release of RB from the albumin nanoparticles was very slow in PBS and dramatically accelerated in the presence of trypsin. Compared with unmodified nanoparticles, the slower release of RB from the surface-modified HSA nanoparticles in the presence of the enzyme suggested that the existence of a steric hydrophilic barrier on the surface of the nanoparticles made digestion of the nanoparticles more difficult.

Administration, Oral↗
PubMed2001

Genomic sequence and expression analyses of human chromatin assembly factor 1 p150 gene.

Chromatin assembly factor-1 (CAF-1) plays essential roles in eukaryotic chromatin assembly during DNA replication (Smith and Stillman, 1989. Cell 58, 15-25), (Krude, 1999. Eur. J. Biochem. 263, 1-5). Its p150 subunit, involved in interaction with histone H3 and H4, is critical to the CAF-1 nucleosome assembly activity. In this study, we sequenced a 96-kb genomic DNA region that includes a 42.8-kb CAF-1 p150 subunit gene (CHAF1A), and a 41.1-kb EEN gene. A scripted bioinformatics analysis pipeline (research agent) has been set up to annotate the BAC sequence with a set of integrated algorithms. The CAF-1 p150 subunit gene contains 15 exons and 14 introns. The promoter region is characterized by deletional analyses, revealing a potential repressor. Tissue-correlated alternative splicing forms of the transcript was initially identified by EST clustering analysis, then confirmed by RT-PCR which resulted more splicing forms than computational prediction.

3T3 Cells↗
PubMed2001

Identification of the human crooked neck gene by comparative gene identification.

Drosophila melanogaster serves as a useful model organism for functional genomic studies, and its genome project was recently completed. We previously described a comparative-gene-identification approach to assist human ortholog gene identification that involves applying an entire proteome as an alignment template. Analysis of the available 14100 Drosophila protein sequences revealed that 37% of them (5228 sequences) might lead to discoveries of novel human genes. Upon further database interrogations, we found several putative full-length human gene transcripts, including the human crooked neck (crn) gene. Based on sequence gap-closure experiments using reverse transcriptase-polymerase chain reaction as well as bioinformatic analysis, we found that the assembled human cDNA contig of crooked neck gene was at least 3903 base pairs in length with alternative splicing variations which encoded mainly for a 687-amino-acid residue protein. The human crooked neck gene was located on chromosome 20 with at least 15 exons. The unique features of the 16 copies of the tetratrico peptide repeat (TPR) motif were conserved in the yeast, fly and human crooked neck orthologous proteins, which were important for spliceosome assembly in cells.

Amino Acid Sequence↗
PubMed2001

A kinetic study on the interaction of deprotonated purine radical cations with amino acids and model peptides.

By use of pulse radiolysis techniques, the radical cations of purine nucleotides have been successfully produced by the SO4- ion oxidation. Time-resolved spectroscopic evidence is provided that the one-electron-oxidized radicals of dAMP and dGMP can be efficiently repaired by aromatic amino acids (including tyrosine and tryptophan) via electron transfer reaction. As a model peptide, Arg-Tyr-AcOH was also investigated with regard to its interaction with deprotonated purine radical cations. The rate constants of the electron transfer reactions were determined to be (1 approximately 5) x 10(8) dm(3) mol(-1) s(-1). These results suggest that the aromatic amino acids in DNA-associated proteins may play some role in electron transfer reactions through DNA.

Amino Acids↗
PubMed2001

Quantitative measurements of cerebral blood flow in patients with unilateral carotid artery occlusion: a PET and MR study.

Although it has been demonstrated that quantitative measures of cerebral blood flow (CBF) can be obtained with the singular value decomposition (SVD) algorithm, the extent to which quantitative CBF measurements can be utilized under pathophysiological conditions has not been systematically studied. A total of five healthy volunteers and five patients with unilateral carotid artery occlusion were studied. Only magnetic resonance (MR) images were acquired for the volunteer group while both MR and positron emission tomography (PET) images were acquired for the patient group. Assessments of CBF from normal volunteers compared favorably with values reported in the literature. However, while a linear relationship was observed for each patient when MR measured CBF was compared to that obtained from PET, this linear relationship diminished when all patients were analyzed as a group (r = 0.41). A correction factor (CF) was proposed that was equal to the ratio of the area of the venous output function (VOF) in each patient to the mean VOF obtained from the volunteer group. After globally scaling the CBF of each patient based on the experimentally derived CF, a substantial improvement was observed (a slope of 1.02 and r = 0.8 for the linear regression line) in the relationship between MR estimated CBF and those obtained from PET.

Analysis of Variance↗
PubMed2001

Lactic acid-stabilised albumin for microsphere formulation and biomedical coatings.

Microspheres of ovalbumin (OVA) ranging from 1 to 15 microm were prepared by emulsifying an aqueous solution of albumin in soya oil at room temperature then raising the temperature to 45 degrees C for 30 min, prior to harvesting of the microspheres. Production of OVA nanospheres with size less than 500 nm was achieved by desolvation from aqueous albumin solutions using acetone. In both cases, lactic acid was added to the starting albumin solution to stabilise the resulting particles. Utilisation of an endogenous substance avoids the use of chemical crosslinking agents such as glutaraldehyde and associated toxicological concerns. Protein coating of knitted Dacron vascular grafts was performed by impregnation of the textile structure with lactic acid-stabilised ovalbumin nanospheres thereby providing a surface potentially resistant to blood platelet adhesion but conducive to endothelialisation. Protein release testing carried out in PBS at 37 degrees C revealed that approximately 60% of the original albumin coating was retained by the Dacron graft material after 4 days and remained at this level for upto 4 weeks. Apart from the formulation of albumin microspheres for drug delivery, diagnostic applications and coating of biomedical textiles, the process of albumin stabilisation using lactic acid may be usefully applied to improve protein immobilisation on a wide range of biomaterial surfaces.

Blood Vessel Prosthesis↗
PubMed2001

Nitric oxide production by coelomocytes of Asterias forbesi.

Vertebrate mononuclear phagocytes produce a plethora of molecules involved in host defense. Among the most potent are the reactive oxygen and nitrogen intermediates. Coelomocytes from invertebrates subserve many of the same functions. In order to determine whether invertebrate phagocytes employ reactive nitrogen intermediates, we investigated the effect of various nonspecific stimulators and invertebrate interleukin (IL)-1alpha- and beta-like molecules on nitric oxide (NO) production. Elevated NO release by stimulated coelomocytes was seen after 24 h. Incubation of stimulated coelomocytes in the presence of arginine analogs inhibited NO release. When invertebrate IL-1-like molecules were added to the coelomocytes, they stimulated the release of NO. Western blot analysis using a polyclonal rabbit antiserum to murine NO synthase detected a band at approximately 125 kDa. These data indicate that coelomocytes are capable of producing and releasing NO and that NO is a chemical mediator that has been conserved as a host defense weapon of phagocytes through evolutionary time.

Animals↗
PubMed2001

Unexpected effects of FERM domain mutations on catalytic activity of Jak3: structural implication for Janus kinases.

Janus kinases comprise carboxyterminal kinase, pseudokinase, SH2-like, and N-terminal FERM domains. We identified three patient-derived mutations in the FERM domain of Jak3 and investigated the functional consequences of these mutations. These mutations inhibited receptor binding and also abrogated kinase activity, suggesting interactions between the FERM and kinase domains. In fact, the domains were found to physically associate, and coexpression of the FERM domain enhanced activity of the isolated kinase domain. Conversely, staurosporine, which alters kinase domain structure, disrupted receptor binding, even though the catalytic activity of Jak3 is dispensable for receptor binding. Thus, the Jak FERM domain appears to have two critical functions: receptor interaction and maintenance of kinase integrity.

Adenosine Triphosphate↗
PubMed2001

Relationship of ultrafiltration and anastomotic flow in isolated rat lungs.

OBJECTIVE: When arterial and venous pressures are increased to equal values in "stop-flow" studies, perfusate continues to enter the pulmonary vasculature from the arterial and venous reservoirs. Losses of fluid from the pulmonary vasculature are due to ultrafiltration and flow through disrupted anastomotic (bronchial) vessels. This study compared the relative sites of ultrafiltration and anastomotic flows at low and high intravascular pressures. METHODS: Isolated rat lungs were perfused for 10 minutes with FITC-dextran, which was used to detect ultrafiltration. Arterial and venous catheters were then connected to reservoirs containing radioactively labeled dextrans at 20 or 30 cm H2O for 10 minutes. The vasculature was subsequently flushed into serial vials, and ultrafiltration and vascular filling during the equal-pressure interval were calculated. RESULTS: Ultrafiltration equaled 0.43 +/- 0.11 mL at 20 cm H2O and was similar to the volume of fresh arterial and venous perfusate which entered and remained in the pulmonary vasculature during the equal-pressure interval (0.45 +/- 0.10 mL). At 30 cm H2O, 0.80 +/- 0.23 mL entered and remained in the vasculature during the equal-pressure interval, replacing the original perfusate, and calculated transudation (0.56 +/- 0.09 mL) was not significantly more than at 20 cm H2O. Fluid also entered the airspaces at 30 cm H2O but not at 20 cm H2O. CONCLUSIONS: At 20 cm H2O, flow through anastomotic vessels occurs at sites that are at the arterial and venous ends of the microcirculation. Flow in exchange vessels remains minimal, permitting measurements of ultrafiltration and exchange. Losses of perfusate from the pulmonary vessels complicate measurements of ultrafiltration at 30 cm H2O.

Animals↗
PubMed2001

Differing processing requirements of four recombinant antigens containing a single defined T-cell epitope for presentation by major histocompatibility complex class II.

A set of predictive rules governing the likelihood of generating a particular peptide-major histocompatibility complex (MHC) class II complex from an intact antigen has not been fully elucidated. We investigated the influence of positional and structural constraints in the region of the epitope by designing a set of recombinant antigens that each contained the well-characterized T-cell epitope moth cytochrome c (MCC) (88-103), which is specifically recognized by the monoclonal antibody (mAb) D4 when complexed with H-2Ek. Our model antigens contained MCC(88-103) either peripherally, at or towards the C-terminus, or internally. Their abilities to bind directly to soluble H-2Ek, and the extent of D4 epitope formation from them by antigen processing-competent and -incompetent cell lines, were determined. Here we report that three of these four antigens yielded MCC(88-103)/H-2Ek complexes independently of the conventional MHC class II antigen-processing and presentation pathway, and in each case the epitope was carried peripherally; two bound directly as intact proteins, probably as a result of spatial separation of the epitope from the major globular domain, and one was processed to peptide by a cell-surface protease. One protein, which carried the epitope inserted into an internal loop, acted as a conventional processing-dependent MCC(88-103) delivery vehicle. Thus, this epitope has different presentation requirements depending on its context. These antigens constitute a panel whose framework could be modified to further define predictive rules for antigen processing for presentation through the different MHC class II complex-generating pathways.

Animals↗
PubMed2001

Frontal lobe dementia with novel tauopathy: sporadic multiple system tauopathy with dementia.

We present a novel tauopathy in a patient with a 10-yr history of progressive frontal lobe dementia and a negative family history. Autopsy revealed mild atrophy of frontal and parietal lobes and severe atrophy of the temporal lobes. There were occasional filamentous tau-positive inclusions, but more interesting were numerous distinctive globular neuronal and glial tau-positive inclusions in both gray and white matter of the neocortex. Affected subcortical regions included substantia nigra, globus pallidus, subthalamic nucleus, and cerebellar dentate nucleus, in a distribution similar to progressive supranuclear palsy (PSP), but without significant accompanying neuronal loss or gliosis. Predominantly straight filaments were detected by electron microscopy (EM), while other inclusions were similar to fingerprint bodies. No twisted ribbons were detected. Immuno-EM studies revealed that only the filamentous inclusions were composed of tau. Immunoblotting of sarkosyl-insoluble tau revealed 2 major bands of 64 and 68 kDa. Blotting analysis after dephosphorylation revealed predominantly 4-repeat tau. Sequence analysis of tau revealed that there were no mutations in either exons 9-13 or the adjacent intronic sequences. The unique cortical tau pathology in this case of sporadic multiple system tauopathy with dementia adds a new pathologic profile to the spectrum of tauopathies.

Aged↗
PubMed2001

Expression and function of the HSD-3.8 gene encoding a testis-specific protein.

The nucleotide sequence of the full length HSD-3.8 cDNA (accession number AF311312), encoding a human sperm component, was determined to consist of 3818 bp with a reading frame of 2778 bp encoding a deduced polypeptide composed of 926 amino acids. A 0.7 kb fragment containing three immunological epitopes of HSD-3.8 cDNA was prepared and used to construct recombinant expression vectors. The constructs were transformed into E.coli BL-21, and the fusion proteins were expressed, isolated and purified. Using the polyclonal antibodies raised against the purified expressed fusion proteins, positive immunostaining occurred over the surface of the postacrosomal zone of human spermatozoa and of germ cells within the seminiferous epithelium of human testis. Intense staining of large pachytene primary spermatocytes occurred. The capacity of the recombinant protein to reduce fertility as an immunogen in adult female rats was assessed. Immunized animals were infertile or exhibited marked reduction in their fertility. Analysis of the deduced HSD-3.8 polypeptide revealed the presence of a tetratricopeptide repeat (TPR) motif, a P-loop sequence that acts as a binding site for ATP/GTP and phosphorylation sites for PKC, CK2 and cAMP/cGMP-dependent protein kinases. A blot overlay assay with [alpha-(32)P]GTP showed that the polypeptide encoded by the 0.7 kb fragment of HSD-3.8 is a GTP binding protein. It was also shown to possess GTPase activity and to be phosphorylated by PKC in vitro. In conclusion, HSD-3.8 is a GTP binding protein and its activity may be regulated by phosphorylation.

Adenosine Triphosphate↗
PubMed2001

Fast repair of the radical cations of dCMP and poly C by quercetin and rutin.

The effects of quercetin and rutin on the repair of the radical cations of dCMP and poly C were studied using the technique of pulse radiolysis. The radical cations of dCMP and poly C were formed by the reaction of dCMP and poly C with SO( 4)(-). After pulse irradiation of nitrogen-saturated aqueous solutions containing dCMP, 20 mM K(2)S(2)O(8), 200 mM t-BuOH and either rutin or quercetin, the initially formed radical cation of dCMP, detected spectrophotometrically, rapidly decayed with the concurrent formation of the phenoxyl radical of rutin or quercetin within 8-40 micros. The repair efficiencies of the tested compounds towards the poly C radical cation were also determined using the same procedure. The results indicate that dCMP and poly C radical cations can be rapidly repaired by quercetin and rutin. The rate constants of the repair reactions were determined to be 4.3-8.8x10(8) M/s and 1.5-3.6x10(8) M/s for dCMP and poly C radical cations, respectively. Together with findings from our previous studies, the present results demonstrate that non-enzymatic fast repair may be a universal form of repair involving phenolic antioxidants.

Antioxidants↗