Biomedical subjects

M Lin

Publications and source records attributed to M Lin.

At least 37 records · Page 2Linked to original sources

PubMed2001

Actin polymerisation during morphogenesis of the acrosome as spermatozoa undergo epididymal maturation in the tammar wallaby (Macropus eugenii).

In the tammar wallaby (Macropus eugenii), post-testicular acrosomal shaping involves a complex infolding and fusion of the anterior and lateral projections of the scoop-shaped acrosome into a compact button-like structure occupying the depression on the anterior end of the sperm nucleus. The present study has generated cytochemical and histological evidence to demonstrate that the occurrence of actin filaments (F-actin, labelled by Phalloidin-FITC) in the acrosome of tammar wallaby spermatozoa is temporally and spatially associated with the process of acrosomal shaping in the epididymis, through a pool of monomeric actin (G-actin, labelled by Rh-DNase I) present in the acrosome throughout all stages of epididymal maturation. F-actin was not detected in the acrosome of testicular spermatozoa, but was found in the infolding and condensing acrosome of caput and corpus epididymal spermatozoa. When the spermatozoa completed acrosome shaping in the cauda epididymidis, F-actin disappeared from the acrosomal area. The strong correlation between the occurrence of F-actin and the events of acrosomal shaping suggested that the post-testicular shaping of the acrosome might depend on a precise succession of assembly and disassembly of F-actin within the acrosome as the spermatozoa transit the epididymis. Thus, actin filaments might play a significant role in the acrosomal transformation, as they are commonly involved in morphological changes in somatic cells.

Acrosome↗

PubMed2001

Polymorphism and distribution of the Secretor alpha(1,2)-fucosyltransferase gene in various Taiwanese populations.

BACKGROUND: The Secretor gene (Se or FUT2), which produces alpha(1,2)-fucosyltransferase, exhibits extensive polymorphism. Six Se genes, including the weak Se (Se(w) or Se385) and three nonsecretor alleles (se571, se685, and se849) have been detected in various populations of Taiwan. The distribution of various Lewis phenotypes among the Taiwanese population groups has been shown to vary considerably. STUDY DESIGN AND METHODS: A PCR-RFLP analysis system, which was based on the nucleotide polymorphism variation of the different Se alleles and which can reveal the Se genotype of an individual easily and accurately, was developed. The distribution of the Se alleles among Taiwanese population groups, including the two major populations, Minnan and Hakka Taiwanese, and 11 indigenous groups, was analyzed by the method. RESULTS: Frequencies of the Se alleles among the Taiwanese populations were revealed. The distribution of the nonsecretor alleles, especially se849, showed a marked variation. A good correlation was observed between a person's Se genotype and Lewis phenotype. CONCLUSION: The Se genes have a polymorphic distribution among various Taiwanese populations, and this agrees with previous results for Lewis phenotype distributions. The Se(w) allele and the three se alleles are responsible for the Le(a+b+) and Le(a+b-) phenotypes, respectively.

Fucosyltransferases↗

PubMed2001

Development of a competitive ELISA using a truncated E2 recombinant protein as antigen for detection of antibodies to classical swine fever virus.

The sequence encoding a truncated E2 glycoprotein of the Alfort/187 strain of classical swine fever virus (CSFV) was expressed in Escherichia coli using the pET expression system and the recombinant product purified by Ni-NTA agarose affinity chromatography. The antigenicity of this recombinant protein was demonstrated by immunoblot using anti- CSFV-specific antibodies. A monoclonal antibody was produced against the truncated E2 protein and used as competitor in an ELISA for the detection of antibodies to CSFV. Specific antibodies were demonstrated by competitive ELISA (C-ELISA) as early as 21 days post-infection (dpi) in experimentally infected pigs. Seroconversion was demonstrated by C-ELISA and neutralising peroxidase-linked assay (NPLA) in all infected animals by 4 weeks. No cross-reaction with antibodies to bovine viral diarrhoea virus (BVDV) was seen in the C-ELISA using sera from experimentally infected pigs. The C-ELISA is not intended as a substitute for the NPLA. However, it is expected it will be useful for monitoring and prevalence studies. It will also assist in testing a large number of samples in the event of an outbreak.

Animals↗

PubMed2001

Three new stilbene trimers from the lianas of Gnetum hainanense.

Three new stilbene trimers, gnetuhainins M-O (1-3), were isolated from the lianas of Gnetum hainanense. Their structures and relative configurations were determined by spectroscopic evidence, especially on 2D NMR analysis. The anti-inflammatory activity has been tested for the isolated compounds.

Molecular Structure↗

PubMed2001

Four new stilbene dimers from the lianas of Gnetum hainanense.

Four new stilbene dimers, gnetuhainins P (1), Q (2), K (3) and L (4), were isolated from the lianas of Gnetum hainanense C. Y. Cheng. Their structures and relative configurations were determined on the basis of spectroscopic evidence, especially 2D NMR techniques.

China↗

PubMed2001

How is the liver primed or sensitized for alcoholic liver disease?

This article represents the proceedings of a symposium at the 2000 ISBRA Meeting in Yokohama, Japan. The chairs were Hidekazu Tsukamoto and Yoshiyuki Takei. The presentations were (1) Tribute to Professor Rajendar K. Chawla, by Craig J. McClain; (2) Dysregulated TNF signaling in alcoholic liver disease, by Craig J. McClain, S. Joshi-Barve, D. Hill, J Schmidt, I. Deaciuc, and S. Barve; (3) The role of mitochondria in ethanol-mediated sensitization of the liver, by Anna Colell, Carmen Garcia-Ruiz, Neil Kaplowitz, and Jose C. Fernandez-Checa; (4) A peroxisome proliferator (bezafibrate) can prevent superoxide anion release into hepatic sinusoid after acute ethanol administration, by Hirokazu Yokoyama, Yukishige Okamura, Yuji Nakamura, and Hiromasa Ishii; (5) S-adenosylmethionine affects tumor necrosis factor-alpha gene expression in macrophages, by Rajendar K. Chawla, S. Barve, S. Joshi-Barve, W. Watson, W. Nelson, and C. McClain; (6) Iron, retinoic acid and hepatic macrophage TNFalpha gene expression in ALD, by Hidekazu Tsukamoto, Min Lin, Mitsuru Ohata, and Kenta Motomura; and (7) Role of Kupffer cells and gut-derived endotoxin in alcoholic liver injury, by N. Enomoto, K. Ikejima, T. Kitamura, H. Oide, Y. Takei, M. Hirose, B. U. Bradford, C. A. Rivera, H. Kono, S. Peter, S. Yamashina, A. Konno, M. Ishikawa, H. Shimizu, N. Sato, and R. Thurman.

Animals↗

PubMed2001

Ionic mechanisms underlying burst firing of layer III sensorimotor cortical neurons of the cat: an in vitro slice study.

We examined the ionic mechanisms underlying burst firing in layer III neurons from cat sensorimotor cortex by intracellular recording in a brain slice. Regular spiking was observed in 77.4% of 137 neurons in response to constant intracellular current pulses of 0.5- to 1-s duration. The rest of the neurons showed burst firing. An initial burst followed by regular-spike firing was seen in 71.0% of 31 bursting neurons. The rest of the bursting neurons (n = 9) exhibited repetitive bursting. In the bursting neurons, spikes comprising the burst were triggered from the afterdepolarization (ADP) of the first spike of the burst. We examined the ionic mechanisms underlying the ADP by applying channel-blocking agents. The ADP was enhanced (rather than blocked) by Ca2+ channel blockade. This enhancement of the ADP by Ca2+ channel blockade was apparent even after blockade of the afterhyperpolarization by apamin or intracellular Ca2+ chelation by EGTA. The firing rate of the regular-spiking cells was increased by apamin, intracellular EGTA or Ca2+ channel blockers. In 17.9% of the neurons examined (n = 56), these agents switched the regular-spiking pattern into a bursting one. Burst firing could not be changed to regular spiking by these agents. Four neurons that responded with a single initial burst in control solution responded with repetitive bursting after application of these agents. We conclude that the main function of Ca2+ influx in layer III neurons is to activate Ca2+-dependent K+ conductance, which prevents or limits burst firing. At a time when spike amplitude was unchanged, the ADP was blocked and the burst firing changed to regular spiking by extracellularly applied tetrodotoxin (TTX) or intracellularly applied N-(2,6-dimethylphenylcarbamoylmethyl) triethyl ammonium bromide (QX314). We concluded that a TTX- and QX314-sensitive Na+ current underlies the ADP and therefore contributes to the burst firing of layer III neurons from the cat cortex.

Action Potentials↗

PubMed2001

Application of competitive enzyme-linked immunosorbent assay for the serologic diagnosis of classical swine fever virus infection.

A competitive enzyme-linked immunosorbent assay (C-ELISA), based on a truncated E2 recombinant protein of the Alfort/187 strain of classical swine fever virus (CSFV) and a specific monoclonal antibody M1669, was evaluated using 2,000 sera from clinically healthy pigs in Canada (a CSFV-free country) and sera from experimentally infected pigs. The relative specificity and sensitivity of the C-ELISA were 100% and 86%, respectively, at a cutoff of 25% inhibition using negative and positive pig sera, as defined by the neutralizing peroxidase-linked assay (NPLA). A kappa value of 0.91 was obtained, indicating an excellent level of agreement between the NPLA and the C-ELISA. When sera from 120 infected pigs were used in the test at > or = 21 days postinfection, the sensitivity of the C-ELISA and the kappa value increased to 97% and 0.98, respectively. This C-ELISA will be useful when a large number of samples must be tested, as could occur during a disease outbreak or for surveillance or prevalence studies.

Animals↗

PubMed2001

Antioxidative activity of natural isorhapontigenin.

Isorhapontigenin (ISOR), isolated from Belamcanda chinensis, is a derivative of stilbene. Its chemical structure is very similar to that of resveratrol, with a potent antioxidative effect. In the present study, we investigated the antioxidative activity of ISOR in vitro. Oxidative damage of rat liver microsomes, brain mitochondria and synaptosomes was induced by Fe2+-Cys, VitC-ADP-Fe2+ and H2O2, respectively. The formation of malondialdehyde (MDA), decrease of reduced glutathione (GSH) and increase of ultra-weak chemiluminescence during the lipid peroxidation process were determined. In addition, the characteristic ultra-weak chemiluminescence of oxidative DNA damage induced by CuSO4-Phen-VitC-H2O2 system was studied. The results showed that ISOR significantly inhibited MDA formation in liver microsomes, brain mitochondria and synaptosomes induced by Fe2+-Cys. Also, ISOR markedly prevented the decrease of GSH in mitochondria and synaptosomes induced by H2O2 and the increase of ultra-weak chemiluminescence during lipid peroxidation induced by VitC-ADP-Fe2+ as well as oxidative DNA damage induced by CuSO4-Phen-VitC-H2O2. The effects of ISOR at 10(-5) and 10(-6) mol/L on the MDA formation and decrease of GSH were similar to that of the classical antioxidant vitamin E (10(-4) mol/L). It may be concluded that ISOR possessed potent antioxidative activity and was much more potent than vitamin E.

Animals↗

PubMed2001

Biochemical and functional analysis of a conserved IGF-binding protein isolated from rainbow trout (Oncorhynchus mykiss) hepatoma cells.

Rainbow trout (Oncorhynchus mykiss) serum contains several IGF-binding proteins (IGFBPs) that specifically bind to IGFs. The structures of these fish IGFBPs have not been determined and their physiological functions are poorly defined. In this study, we identified a 30 kDa IGFBP present in rainbow trout serum and secreted by cultured trout hepatoma cells. This IGFBP binds to IGFs but not to insulin. This IGFBP was purified to homogeneity using a three-step procedure involving Phenyl-Sepharose chromatography, IGF-I affinity chromatography and reverse-phase HPLC. Affinity cross-linking studies indicated that this IGFBP binds to IGF-I with a higher affinity than to IGF-II. N-terminal sequence analysis of the trout IGFBP suggests that it shares high sequence identity with that of human IGFBP-1 in the N-terminal region. When added to cultured fish and human cells, the trout IGFBP inhibited IGF-I-stimulated DNA synthesis and cell proliferation in a concentration-dependent manner. The inhibitory effect of the fish IGFBP was comparable to those of human IGFBP-1 and -4. These results indicate that the IGFBP molecule is structurally and functionally conserved in evolutionarily ancient vertebrate species such as bony fish.

Amino Acid Sequence↗

PubMed2000

Molecular basis of the Kell-null phenotype: a mutation at the splice site of human KEL gene abolishes the expression of Kell blood group antigens.

The Kell blood group system is polymorphic, and 23 antigens have been defined to date. The Kell antigens are located on a single red cell transmembrane glycoprotein, encoded by the 19 exons of the KEL gene. The different Kell phenotypes result from point mutations leading to amino acid changes in the Kell glycoprotein. An unusual phenotype, which is defined as the complete lack of all of the Kell antigens, has been identified and designated as the Kell-null or Ko phenotype. The coding region of the KEL gene of the Ko individual showed a normal KEL2/KEL4/KEL7 gene sequence; nevertheless, a G to C mutation at the splice donor site (5' splice site) of intron 3 was found to be present as a homozygote in the individual. The mutation destroys the conserved GT sequence of the splice donor site. Reverse transcription-polymerase chain reaction analysis showed the absence of the complete KEL mRNA. Instead, a major transcript with the exon 3 region skipped was found. The exon 3 of the KEL gene encodes the transmembrane domain of the Kell glycoprotein, and a transcript without exon 3 is predicted to have a premature stop codon that abolishes the translation of C-terminal segment. The segment contains all of the known positions responsible for characterizing different Kell antigens, and this explains the lack of all Kell antigens in Ko red cells.

Alleles↗

PubMed2000

cDNA cloning and expression of a human aldehyde dehydrogenase (ALDH) active with 9-cis-retinal and identification of a rat ortholog, ALDH12.

This report describes the isolation of a heretofore uncharacterized aldehyde dehydrogenase (ALDH) with retinal dehydrogenase activity from rat kidney and the cloning and expression of a cDNA that encodes its human ortholog, the previously unknown ALDH12. The human ALDH12 cDNA predicts a 487-residue protein with the 23 invariant amino acids, four conserved regions, cofactor binding motif (G(209)XGX(3)G), and active site cysteine residue (Cys(287)) that typify members of the ALDH superfamily. ALDH12 seems at least as efficient (V(m)/K(m)) in converting 9-cis-retinal into the retinoid X receptor ligand 9-cis-retinoic acid as two previously identified ALDHs with 9-cis-retinal dehydrogenase activity, rat retinal dehydrogenase (RALDH) 1 and RALDH2. ALDH12, however, has approximately 40-fold higher activity with 9-cis- retinal than with all-trans-retinal, whereas RALDH1 and RALDH2 have equivalent and approximately 4-fold less efficiencies for 9-cis-retinal versus all-trans-retinal, respectively. Therefore, ALDH12 is the first known ALDH to show a preference for 9-cis-retinal relative to all-trans-retinal. Evidence consistent with the possibility that ALDH12 could function in a pathway of 9-cis-retinoic acid biosynthesis in vivo includes biosynthesis of 9-cis-retinoic acid from 9-cis-retinol in cells co-transfected with cDNAs encoding ALDH12 and the 9-cis-retinol/androgen dehydrogenase, cis-retinoid/androgen dehydrogenase type 1. Intense ALDH12 mRNA expression in adult and fetal liver and kidney, two organs that reportedly have relatively high concentrations of 9-cis-retinol, reinforces this notion.

Aldehyde Dehydrogenase↗

PubMed2000

Functional roles of charged residues in the putative voltage sensor of the HCN2 pacemaker channel.

Hyperpolarization-activated, cyclic nucleotide-gated (HCN) channels contribute to pacemaking activity in specialized neurons and cardiac myocytes. HCN channels have a structure similar to voltage-gated K(+) channels but have a much larger putative S4 transmembrane domain and open in response to membrane hyperpolarization instead of depolarization. As an initial attempt to define the structural basis of HCN channel gating, we have characterized the functional roles of the charged residues in the S2, S3, and S4 transmembrane domains. The nine basic residues and a single Ser in S4 were mutated individually to Gln, and the function of mutant channels was analyzed in Xenopus oocytes using two-microelectrode voltage clamp techniques. Surface membrane expression of hemagglutinin-epitope-tagged channel proteins was examined by chemiluminescence. Our results suggest that 1) Lys-291, Arg-294, Arg-297, and Arg-300 contribute to the voltage dependence of gating but not to channel folding or trafficking to the surface membrane; 2) Lys-303 and Ser-306 are essential for gating, but not for channel folding/trafficking; 3) Arg-312 is important for folding but not gating; and 4) Arg-309, Arg-315, and Arg-318 are crucial for normal protein folding/trafficking and may charge-pair with Asp residues located in the S2 and S3 domains.

Amino Acid Sequence↗

PubMed2000

A structural basis for drug-induced long QT syndrome.

Mutations in the HERG K(+) channel gene cause inherited long QT syndrome (LQT), a disorder of cardiac repolarization that predisposes affected individuals to lethal arrhythmias [Curran, M. E. , Splawski, I., Timothy, K. W., Vincent, G. M., Green, E. D. & Keating, M. T. (1995) Cell 80, 795-804]. Acquired LQT is far more common and is most often caused by block of cardiac HERG K(+) channels by commonly used medications [Roden, D. M., Lazzara, R., Rosen, M., Schwartz, P. J., Towbin, J. & Vincent, G. M. (1996) Circulation 94, 1996-2012]. It is unclear why so many structurally diverse compounds block HERG channels, but this undesirable side effect now is recognized as a major hurdle in the development of new and safe drugs. Here we use alanine-scanning mutagenesis to determine the structural basis for high-affinity drug block of HERG channels by MK-499, a methanesulfonanilide antiarrhythmic drug. The binding site, corroborated with homology modeling, is comprised of amino acids located on the S6 transmembrane domain (G648, Y652, and F656) and pore helix (T623 and V625) of the HERG channel subunit that face the cavity of the channel. Other compounds that are structurally unrelated to MK-499, but cause LQT, also were studied. The antihistamine terfenadine and a gastrointestinal prokinetic drug, cisapride, interact with Y652 and F656, but not with V625. The aromatic residues of the S6 domain that interact with these drugs (Y652 and F656) are unique to eag/erg K(+) channels. Other voltage-gated K(+) (Kv) channels have Ile and Val (Ile) in the equivalent positions. These findings suggest a possible structural explanation for how so many commonly used medications block HERG but not other Kv channels and should facilitate the rational design of drugs devoid of HERG channel binding activity.

Amino Acid Sequence↗

PubMed2000

Cholesteryl ester transfer protein and phospholipid transfer protein have nonoverlapping functions in vivo.

Plasma phospholipid transfer protein (PLTP) and cholesteryl ester transfer protein (CETP) are homologous molecules that mediate neutral lipid and phospholipid exchange between plasma lipoproteins. Biochemical experiments suggest that only CETP can transfer neutral lipids but that there could be overlap in the ability of PLTP and CETP to transfer or exchange phospholipids. Recently developed PLTP gene knock-out (PLTP0) mice have complete deficiency of plasma phospholipid transfer activity and markedly reduced high density lipoprotein (HDL) levels. To see whether CETP can compensate for PLTP deficiency in vivo, we bred the CETP transgene (CETPTg) into the PLTP0 background. Using an in vivo assay to measure the transfer of [(3)H]PC from VLDL into HDL or an in vitro assay that determined [(3)H]PC transfer from vesicles into HDL, we could detect no phospholipid transfer activity in either PLTP0 or CETPTg/PLTP0 mice. On a chow diet, HDL-PL, HDL-CE, and HDL-apolipoprotein AI in CETPTg/PLTP0 mice were significantly lower than in PLTP0 mice (45 +/- 7 versus 79 +/- 9 mg/dl; 9 +/- 2 versus 16 +/- 5 mg/dl; and 51 +/- 6 versus 100 +/- 9, arbitrary units, respectively). Similar results were obtained on a high fat, high cholesterol diet. These results indicate 1) that there is no redundancy in function of PLTP and CETP in vivo and 2) that the combination of the CETP transgene with PLTP deficiency results in an additive lowering of HDL levels, suggesting that the phenotype of a human PLTP deficiency state would include reduced HDL levels.

Animals↗

PubMed2000

Human histo-blood group ABO glycosyltransferase genes: different enhancer structures with different transcriptional activities.

The enhancer element of the human histo-blood group ABO glycosyltransferase gene has been demonstrated to be located -3.7 kb upstream from the transcription start site and to be composed of four tandem repeats of a 43-bp unit. Recently we identified three different enhancer structures among the allelic A, B, and O glycosyltransferase genes. The enhancer structure with four 43-bp units is present in the B and O genes, but not in the A gene. The corresponding enhancer region of the A gene contains only one 43-bp unit, and within this unit a nucleotide substitution exists when compared with the consensus sequence. Through transient transfection assays, the transcriptional activity of the A-gene enhancer region was demonstrated to be less than 1% of that of the B-gene enhancer. The difference between the transcriptional activities of the two enhancers became more significant when acting in concert with the ABO-gene's native promoter. The different repeat numbers of the 43-bp unit possessed by the two allelic genes were shown to be the main reason for the vast difference in the transcriptional activities between the A-gene and B-gene enhancers.

ABO Blood-Group System↗

PubMed2000

Expression of human prostatic acid phosphatase gene is regulated by upstream negative and positive elements.

Human prostatic acid phosphatase (PAcP) is a prostate epithelium-specific differentiation antigen. To understand the regulation of expression of the PAcP gene, we studied the cis-regulatory elements of its promoter. A DNA fragment from -2899 to +87 base pairs (bp) of PAcP gene was fused to the chloramphenicol acetyltransferase (CAT) reporter gene and introduced into PC-3 and LNCaP human prostate cancer cells. The expression of the CAT gene driven by the PAcP promoter was assessed in transient expression assays. Sequential 5' deletions of the promoter were constructed and analyzed to reveal the positive and the negative regulatory elements that are involved in regulating the transcription of the PAcP gene. Our data showed that the proximal sequence -1305/+87 bp directs a high level of the CAT activity in both cell lines. Deletion of the region from -1305 to -779 resulted in approximately a 10- and three-fold decrease of the PAcP promoter activity in PC-3 and LNCaP cells, respectively. Interestingly, an inverse correlation of the CAT activity with the cell growth was observed when the reporter gene was driven by the -1305/+87 fragment, but not by the -779/+87 fragment. Two regions of transcriptional suppression were identified and located in positions from -2899 to -2583, and from -2583 to -1305 bp. Furthermore, the activity of the core promoter region from -779 to +87 bp can be activated by a SV-40 enhancer. The results, thus, clearly demonstrate the presence of positive and negative cis-elements in the promoter region of the PAcP gene.

Acid Phosphatase↗

PubMed2000

Obstructive sleep apnea syndrome in obese Singapore children.

We set out to determine the prevalence of obstructive sleep apnea syndrome (OSAS) among obese Singapore school children and identify risk factors for OSAS. This study was designed as a prospective study in three phases. Parents completed a questionnaire with regards to sleep and daytime symptoms in Phase 1. Children suspected to have OSAS based on the questionnaire and all with a percent ideal body weight (IBW) >/=180 were called for clinic visits in Phase 2. All whose percent IBW >/=180 and those in whom the physicians strongly suspected OSAS were subjected to a polysomnogram in phase 3. The children were recruited from the School Health Nutritional Clinic for obese children. The investigations were carried out at Tan Tock Seng Hospital. In all, 3,671 children were screened with the questionnaire. Of these, 146 were selected to undergo polysomnography. Twenty-six had abnormal sleep studies with apnea/hypoxia indices (AHIs) >5/hr. The significant clinical feature which correlated with OSAS was sleep sitting up (P = 0.005). The risk is higher in morbidly obese (IBW >/=180), with a prevalence of 13.3% (8/60), than in less obese children (IBW <180). One in eight (12.5%) of these children was asymptomatic and would have been missed based on the questionnaire. Presence of adenotonsillar hypertrophy led to increased risk of OSAS. The prevalence of OSAS was 0.7% (26/3,671) among the obese schoolchildren in Singapore, which is similar to the prevalence reported by others. Using discriminant analysis, the estimated prevalence increased to 5.7%. In the morbidly obese (IBW >/=180), the prevalence rate is higher at 13.3%.

Adenoids↗