Biomedical subjects

X Lin

Publications and source records attributed to X Lin.

At least 289 records · Page 16Linked to original sources

PubMed1997

Acetylcholine, outer hair cell electromotility, and the cochlear amplifier.

The dominant efferent innervation of the cochlea terminates on outer hair cells (OHCs), with acetylcholine (ACh) being its principal neurotransmitter. OHCs respond with a somatic shape change to alterations in their membrane potential, and this electromotile response is believed to provide mechanical feedback to the basilar membrane. We examine the effects of ACh on electromotile responses in isolated OHCs and attempt to deduce the mechanism of ACh action. Axial electromotile amplitude and cell compliance increase in the presence of the ligand. This response occurs with a significantly greater latency than membrane current and potential changes attributable to ACh and is contemporaneous with Ca2+ release from intracellular stores. It is likely that increased axial compliance largely accounts for the increase in motility. The mechanical responses are probably related to a recently demonstrated slow efferent effect. The implications of the present findings related to commonly assumed efferent behavior in vivo are considered.

Acetylcholine↗

PubMed1997

Genomic organization and promoter structure of the human EXT1 gene.

Hereditary predisposition to multiple exostoses is a genetically heterogeneous disease. Recently, we have reported the identification of the EXT1 gene on human chromosome 8. We have now isolated a cDNA clone from a human adult lung cDNA library and have determined the genomic organization and promoter structure of the EXT1 gene. The gene is composed of 11 exons, ranging from 90 to 1735 bp, and spans approximately 350 kb of genomic DNA. Sequence analysis of the promoter region revealed the presence of a CpG island containing GC and CAAT boxes, but no TATA box. Such a promoter is characteristic for housekeeping genes. This finding is in good agreement with the ubiquitous expression of the EXT1 gene.

Adult↗

PubMed1997

Fluorescent probes attached to Cys 35 or Cys 84 in cardiac troponin C are differentially sensitive to Ca(2+)-dependent events in vitro and in situ.

The goal of the current study was to generate recombinant cTnC proteins with single Cys residues as sites for attachment of fluorescent probes that can distinguish between the structural effects of myosin cross bridges and direct Ca2+ binding to cTnC (cardiac and slow skeletal troponin C) in skinned fibers. We anticipated that cTnC proteins which retain the endogenous Cys 35 (cTnC(C35)) or Cys 84 (cTnC(C84)) would provide fluorescent probes with distinct microenvironments, since these residues are on opposite sides of the globular regulatory domain. In vitro experiments that showed IAANS (2-(4'-(iodoacetamido)anilino)naphthalene-6-sulfonic acid) coupled to Cys 35 can induce unwanted structural perturbations as evidenced by a decreased affinity of site II for Ca2+ when IAANS-labeled cTnC(C35) is bound to cTnI. Important structural features involving Cys 35 in the inactive site I are suggested by a Ca(2+)-dependent increase in reactivity of Cys 35 with sulfhydryl specific reagents when cTnC(C35) is associated with cTnI. These characteristics are not seen for cTnC(C84). When incorporated in situ into skinned cardiac muscle fibers, native cTnC with IAANS bound to both Cys 35 and Cys 84 showed a pCa50 of fluorescence which preceded that of force, while the pCa50 values of both force and fluorescence were coincident for IAANS-labeled cTnC(C84). Disruption of force-producing myosin cross bridges had no effect on the pCa50 of fluorescence for IAANS-labeled cTnC(C84), but induced a rightward shift in the pCa50 of fluorescence for IAANS-labeled native cTnC. These data can be interpreted to indicate that cTnC with IAANS bound to both Cys 35 and C84 senses either myosin cross bridges or direct Ca2+ binding and myosin-induced cooperativity, while IAANS bound to Cys 84 alone senses conformations that are tightly coupled with force generation.

Amino Acid Sequence↗

PubMed1997

Differential generation of class I H-2D- versus H-2K-restricted cytotoxicity against a demyelinating virus following central nervous system infection.

Despite the fact that both H-2K and D molecules are up-regulated in the central nervous system (CNS) following Theiler's murine encephalomyelitis virus (TMEV) infection, resistance in this virus model of multiple sclerosis maps exclusively to D. To address this paradox, we examined the ability of the K and D molecules to present viral antigens to cytotoxic T lymphocytes (CTL). Whereas no virus-specific CTL were detected in the CNS of susceptible B10.Q and B10.S mice 7 days post-infection, D-restricted CTL were identified readily in the CNS of resistant B10 animals. There was no evidence of K-restricted CTL in the CNS of B10 mice at day 7 post-infection. The presence of both K- and D-restricted virus-specific CTL in the spleen of immunized B10 mice demonstrates that the exclusive use of D molecules by CTL in the CNS of mice 7 days post-infection is not due to the inability of the K molecules to present viral peptides to lymphocytes. We conclude that the prominent role of the D locus in determining resistance or susceptibility to TMEV-induced demyelination is determined by factors governing the regulation of the immune response, and not by the presence or absence of CTL precursors capable of recognizing viral peptides presented by the K and D antigen-presenting molecules, or by differences in the ability of the K and D molecules to present viral peptides.

Animals↗

PubMed1997

Identification of novel mutations in the human EXT1 tumor suppressor gene.

Hereditary multiple exostoses (EXT) is a genetically heterogeneous bone disorder caused by genes segregating on human chromosomes 8, 11, and 19 and designated EXT1, EXT2 and EXT3, respectively. Recently, the EXT1 gene has been isolated and partially characterized and appears to encode a tumor suppressor gene. We have identified six mutations in the human EXT1 gene from six unrelated multiple exostoses families segregating for the EXT gene on chromosome 8. One of the mutations we detected is the same 1-bp deletion in exon 6 that was previously reported in two independent EXT families. The other five mutations, in exons 1, 6, 9, and the splice junction at the 3' end of exon 2, are novel. In each case, the mutation is likely to result in a truncated or nonfunctional EXT1 protein. These results corroborate and extend the previous report of mutations in this gene in two EXT families, and provide additional support for the EXT1 gene as the cause of hereditary multiple exostoses in families showing linkage to chromosome 8.

Alternative Splicing↗

PubMed1997

Relationship of interictal epileptiform discharges to sleep depth in partial epilepsy.

Non-rapid eye movement (NREM) sleep activates interictal epileptiform discharges (spikes) in many epileptic syndromes. To define this phenomenon more precisely, we studied the relationship of spikes to absolute log delta power (LDP), a continuous measure of sleep depth, in 8 patients with partial epilepsy. LDP differed significantly across visually scored sleep stages. Logistic regression analyses of spike occurrence in relation to LDP were carried out on the central-occipital channel contralateral to the dominant spike focus (C4-O2 for left and C3-O1 for right temporal focus). Within NREM sleep, spikes were more likely to occur: (1) at higher levels of LDP, (2) on the ascending limb of LDP, and (3) with more rapid rises in LDP. Spike frequency per minute was 4.6 times higher in NREM than in rapid-eye movement (REM) sleep and diminished with time from sleep onset. When the effect of LDP was controlled for in the analysis, however, there was no significant effect of REM sleep stage or time on spike occurrence. Only 1% of spikes occurred within 10 s of an arousal. These findings suggest that processes underlying the deepening of NREM sleep may contribute to spike activation in partial epilepsy.

Adult↗

PubMed1997

Action potentials and underlying voltage-dependent currents studied in cultured spiral ganglion neurons of the postnatal gerbil.

The excitability of cultured spiral ganglion (SG) neurons from early postnatal gerbil (P0-P1) was examined with the whole-cell patch-clamp technique. The role of voltage-gated currents in shaping the kinetics of action potentials (APs) was analyzed. Cultured SG neurons displayed spontaneous APs with a low rate (< 0.1 Hz). The kinetics of APs were studied by injecting neurons with current pulses of various frequencies and duration. A single depolarizing pulse of long duration elicited only one AP in most SG neurons. When excited by a train of short current pulses given at rates greater than 50 Hz, the firing pattern displayed an adaptive mechanism with the result that successive APs fired with lower amplitude, broader duration and delayed peak time. Pulse trains of higher frequencies had higher failure rates in initiating APs. Current pulses given at 20 Hz or lower elicited APs that had very similar amplitudes. However, the width of the APs gradually broadened. Duration of APs was also found to be affected by the membrane potential of neurons. Between -75 mV and -55 mV, AP duration was broadened at a rate of about 33% per 10 mV depolarization. Voltage-gated currents that underlie the generation of APs were examined under voltage-clamp conditions. Tetrodotoxin-sensitive sodium currents and dihydropyridine-sensitive L-type calcium currents were found. More importantly, inactivation properties of the potassium current provided a direct explanation for the cumulative broadening of APs. This work demonstrated that SG neurons were able to fire APs long before hearing commences in gerbil. Possible roles of spontaneous APs in the development of the cochlea and the role of voltage-gated currents in the function of SG neurons under normal and pathological conditions are discussed.

Action Potentials↗

PubMed1997

Impaired gastric myoelectrical activity in patients with chronic renal failure.

Dysmotility and delayed emptying of the stomach have been reported in patients with chronic renal failure (CRF). The aim of this study was to investigate whether gastric myoelectrical activity was impaired in patients with CRF using electrogastrography. The electrogastrogram (EGG) was recorded in 24 symptomatic patients with CRF (15 with diabetes) and 12 normal subjects. Two 30-min EGG recordings before and after a test meal were analyzed using spectral analysis methods. It was found that patients with CRF showed a significantly lower percentage of normal 2-4 cpm slow waves in both fasting and fed states in comparison with healthy controls (in fasting state: 88.9 +/- 2.5% vs 67.4 +/- 6.6%/63.2 +/- 7.0%, P < 0.01; in fed state: 89.6 +/- 1.8% vs 64.6 +/- 6.2%/62.0 +/- 8.3%, P < 0.01; controls vs diabetic patients/nondiabetic patients). Both patient groups showed a significantly higher prevalence of the abnormal EGG, which was defined as the percentage of 2-4 cpm slow waves lower than 70% (fasting state: 8% vs 60%/56%, P < 0.01/0.05; fed state: 0% vs 53%/56%, P < 0.005/0.002; controls vs diabetic patients/nondiabetic patients). No significant difference was observed in the regularity of the gastric slow waves between the two patient groups. The healthy controls showed a significant increase in the dominant power and frequency of the EGG after the test meal. However, this increase was absent in the two patient groups. It was concluded that patients with chronic renal failure have abnormal gastric myoelectrical activity, including impaired regularity of the gastric slow wave and a failed increase in the power of the EGG at 3 cpm. Electrogastrography is an attractive noninvasive method for the study of gastric motility in patients with severe chronic renal failure.

Adult↗

PubMed1997

Expression of the Bacillus licheniformis PWD-1 keratinase gene in B. subtilis.

The kerA gene which encodes the enzyme keratinase was isolated from the feather-degrading bacterium Bacillus licheniformis PWD-1. The entire gene, including pre-, pro- and mature protein regions, was cloned with Pker, its own promoter, P43, the vegetative growth promoter, or the combination of P43-Pker into plasmid pUB18. Transformation of the protease-deficient strain B. subtilis DB104 with these plasmids generated transformant strains FDB-3, FDB-108 and FDB-29 respectively. All transformants expressed active keratinase in both feather and LB media, in contrast to PWD-1, in which kerA was repressed when grown in LB medium. With P43-Pker upstream of kerA, FDB-29 displayed the highest activity in feather medium. Production of keratinase in PWD-1 and transformants was further characterized when glucose or casamino acids were supplemented into the feather medium. These studies help understand the regulation of kerA expression and, in the long run, can help strain development and medium conditioning for the production of this industrially important keratinase.

Bacillus↗

PubMed1997

Predictors of sleepiness in epilepsy patients.

Sleepiness, a common complaint of epilepsy patients, is frequently attributed to antiepileptic medications. To determine predictors of subjective sleepiness in epilepsy patients, we gave self-administered, validated surveys of sleepiness [Epworth sleepiness scale (our major outcome measure)] and sleep apnea [sleep apnea scale of the sleep disorders questionnaire (SA/SDQ)] to 158 epilepsy patients and 68 neurology patients without epilepsy (controls). An elevated Epworth score (>10) was more likely in epilepsy patients compared to controls after controlling for age and gender (p < 0.05). When Epworth scores were adjusted for SA/SDQ scores and restless legs symptoms (RLS), however, epilepsy patients showed only a nonsignificant trend toward elevated Epworth scores compared to controls (p = 0.08). SA/SDQ scores (p < 0.005) and RLS (p < 0.007) were significant predictors of elevated Epworth score in both epilepsy patients and controls. Among the epilepsy patients, the number or type of antiepileptic medication, seizure frequency, epilepsy syndrome (partial vs. generalized), and the presence of sleep-related seizures were not significant predictors (p > 0.10) of elevated Epworth score. Before attributing sleepiness in epilepsy patients to antiepileptic medications or uncontrolled seizures, clinicians should consider the possibility of a coexisting sleep disorder.

Adolescent↗

PubMed1997

Extensive HLA class II studies in 58 non-DRB1*15 (DR2) narcoleptic patients with cataplexy.

Narcolepsy is a sleep disorder that has been shown to be tightly associated with HLA DR15 (DR2). In this study, 58 non-DR15 patients with narcolepsy-cataplexy were typed at the HLA DRB1, DQA1 and DQB1 loci. Subjects included both sporadic cases and narcoleptic probands from multiplex families. Additional markers studied in the class II region were the promoters of the DQA1 and DQB1 genes, two CA repeat polymorphisms (DQCAR and DQCARII) located between the DQA1 and DQB1 genes, three CA repeat markers (G51152, T16CAR and G411624R) located between DQB1 and DQB3 and polymorphisms at the DQB2 locus. Twenty-one (36%) of these 58 non-DR15 narcoleptic patients were DQA1*0102 and DQB1*0602, a DQ1 subtype normally associated with DRB1*15 in DR2-positive narcoleptic subjects. Additional microsatellite and DQA1 promoter diversity was found in some of these non-DR15 but DQB1*0602-positive haplotypes but the known allele specific codons of DQA1*0102 and DQB1*0602 were maintained in all 21 cases. The 37 non-DQA1*0102/DQB1*0602 subjects did not share any particular HLA DR or DQ alleles. We conclude that HLA DQA1*0102 and DQB1*0602 are the most likely primary candidate susceptibility genes for narcolepsy in the HLA class II region.

Cataplexy↗

PubMed1997

Role of sialyloligosaccharide binding in Theiler's virus persistence.

Theiler's murine encephalomyelitis viruses (TMEVs) belong to the Picornaviridae family and are divided into two groups, typified by strain GDVII virus and members of the TO (Theiler's original) group. The highly virulent GDVII group causes acute encephalitis in mice, while the TO group is less virulent and causes a chronic demyelinating disease which is associated with viral persistence in mice. This persistent central nervous system infection with demyelination resembles multiple sclerosis (MS) in humans and has thus become an important model for studying MS. It has been shown that some of the determinants associated with viral persistence are located on the capsid proteins of the TO group. Structural comparisons of two persistent strains (BeAn and DA) and a highly virulent strain (GDVII) showed that the most significant structural variations between these two groups of viruses are located on the sites that may influence virus binding to cellular receptors. Most animal viruses attach to specific cellular receptors that, in part, determine host range and tissue tropism. In this study, atomic models of TMEV chimeras were built with the known structures of GDVII, BeAn, and DA viruses. Comparisons among the known GDVII, BeAn, and DA structures as well as the predicted models for the TMEV chimeras suggested that a gap on the capsid surface next to the putative receptor binding site, composed of residues from VP1 and VP2, may be important in determining viral persistence by influencing virus attachment to cellular receptors, such as sialyloligosaccharides. Our results showed that sialyllactose, the first three sugar molecules of common oligosaccharides on the surface of mammalian cells, inhibits virus binding to the host cell and infection with the persistent BeAn virus but not the nonpersistent GDVII and chimera 39 viruses.

Amino Acid Sequence↗

PubMed1997

Brain-infiltrating cytolytic T lymphocytes specific for Theiler's virus recognize H2Db molecules complexed with a viral VP2 peptide lacking a consensus anchor residue.

Mice expressing the H2b haplotype are resistant to infection with Theiler's murine encephalomyelitis virus (TMEV), which causes chronic demyelination in susceptible mice. The prominent cytolytic T-lymphocyte (CTL) response to the VP2 antigen encoded by TMEV led us to the identification of a class I-binding peptide derived from the VP2 antigen. Escherichia coli transformants overexpressing a series of 11 overlapping VP2 protein fragments were subjected to lysis and alkali digestion, and the resultant peptide pools were tested for their abilities to sensitize RMA-S targets for lysis by CTLs. The source of effector CD8+ T cells for the assays was either freshly harvested central nervous system-infiltrating lymphocytes (CNS-IL) or CNS-IL-derived VP2-specific CTL clones and lines. A 10-residue peptide at VP2 positions 121 to 130 (VP2(121-130)) (FHAGSLLVFM) was identified that sensitized targets for lysis and formed stable complexes with H2Db class I molecules. The VP2(121-130) peptide sensitized target cells for lysis by freshly harvested CNS-IL CTLs at femtomolar concentrations. Despite its relative high level of biological activity, the VP2(121-130) peptide is distinguished from other Db-binding peptides by its lack of an asparagine residue at position five, which had been previously proposed to be a requirement for Db-peptide complexing.

Amino Acid Sequence↗

PubMed1997

Hepatic fatty acid metabolism in pigs and rats: major differences in endproducts, O2 uptake, and beta-oxidation.

Models of mammalian hepatic lipid metabolism are based largely on observations made in adult rats, emphasizing ketogenesis as a primary adjunct to mitochondrial beta-oxidation. Studies using piglets have illustrated the divergent nature of intermediary metabolism in this model, wherein ketogenesis and beta-oxidation are small and acetogenesis is an important route of fuel carbon flux. To clarify potential species differences in hepatic lipid metabolism and its control, we compared O2 consumption and metabolic end products in fasted pig and rat liver homogenates treated with 1-[14C]C16:0. Carboxyl carbon accumulation in acid-soluble products (ASP) plus CO2 was threefold greater and O2 consumption was twofold greater in rats (P < 0.05). Unlike rats, pigs showed negligible carboxyl carbon accumulation in ketone bodies (3-7% of ASP), whereas acetate was a carboxyl carbon reservoir in both animals (17-31% of ASP in pigs). Malonate increased (approximately 2-fold) and antimycin/rotenone decreased (40-60%) radiolabel accumulation in acetate. These data concur with the hypotheses that comparatively low hepatic beta-oxidative flux in piglets is partially related to a smaller metabolic rate and that substantial acetogenesis occurs intramitochondrially in both pigs and rats.

Animals↗

PubMed1997

Response of hepatic mitochondrial and peroxisomal beta-oxidation to increasing palmitate concentrations in piglets.

Responses of total, mitochondrial, and peroxisomal beta-oxidation to increasing [1-14C]-palmitate concentrations (0.02-1.0 mM) were measured in liver homogenates from neonatal pigs. Incubations were conducted in the absence (total beta-oxidation) or presence (peroxisomal beta-oxidation) of antimycin A and rotenone; mitochondrial beta-oxidation was calculated as total minus peroxisomal oxidation. Total and mitochondrial beta-oxidations were maximized at a palmitate concentration of 0.05 mM, whereas peroxisomal beta-oxidation was maximized at 0.50 mM palmitate. Across concentrations, peroxisomal beta-oxidation contributed 40-47% of total beta-oxidation. An increased rate of CO2 production and a greater ratio of CO2 production to total mitochondrial beta-oxidation as palmitate concentration increased suggested that the limited capacity for mitochondrial beta-oxidation was attributable primarily to limited ketogenic capacity. Comparative observations in liver from adult rats showed that peroxisomal beta-oxidation was maximized at 0.1 mM palmitate, but total and mitochondrial beta-oxidation rates were not maximized even at 1 mM palmitate. At 1 mM palmitate, peroxisomal beta-oxidation was 20% of total beta-oxidation in adult rats and 37% in adult pigs. Therefore, the contribution of peroxisomal beta-oxidation to total beta-oxidation is highly dependent on substrate concentration and appears to be greater in adult pigs than in adult rats. The greater proportional contribution of peroxisomal beta-oxidation in piglet liver might act as a compensatory mechanism for piglets to oxidize milk fatty acids.

Animals↗

PubMed1997

The Drosophila sugarless gene modulates Wingless signaling and encodes an enzyme involved in polysaccharide biosynthesis.

We have identified and characterized a Drosophila gene, which we have named sugarless, that encodes a homologue of vertebrate UDP-glucose dehydrogenase. This enzyme is essential for the biosynthesis of various proteoglycans, and we find that in the absence of both maternal and zygotic activities of this gene, mutant embryos develop with segment polarity phenotypes reminiscent to loss of either Wingless or Hedgehog signaling. To analyze the function of Sugarless in cell-cell interaction processes, we have focused our analysis on its requirement for Wingless signaling in different tissues. We report that sugarless mutations impair signaling by Wingless, suggesting that proteoglycans contribute to the reception of Wingless. We demonstrate that overexpression of Wingless can bypass the requirement for sugarless, suggesting that proteoglycans modulate signaling by Wingless, possibly by limiting its diffusion and thereby facilitating the binding of Wingless to its receptor. We discuss the possibility that tissue-specific regulation of proteoglycans may be involved in regulating both Wingless short- or long-range effects.

Amino Acid Sequence↗

PubMed1997

The mechanism and site of action of lidocaine hydrochloride in guinea pig inner ear.

Lidocaine was applied to the round window (RW) in order to localize its site of action in the cochlea. Cochlear microphonic (CM), summating potential (SP), and compound action potential (CAP) input/output functions were measured to a 16 kHz tone burst to assess the functional changes of the cochlea. In separate experiments, the effect of lidocaine on the whole cell current of isolated outer hair cells (OHC) was studied. A dose of 2 microliters of 40 mM lidocaine in saline solution, when applied to the RW, caused a small change in all measured variables, indicating a passage of the drug through the RW membrane to sites of action. However, 160 mM of lidocaine further decreased CM, SP, and CAP by a total of 40% from the control. A partial recovery occurred for CM during the 30 min follow-up period. CAP and SP continued to decline. In isolated OHCs, lidocaine decreased the whole cell current in a dose-dependent fashion. The KD for lidocaine effect on OHCs was 7 mM. Our in vivo results indicate that lidocaine affects OHCs and reduces CM, causing a subsequent reduction in SP and CAP. The increased effect of lidocaine on CAP and SP, while CM is recovering, suggests an additional specific effect of lidocaine on the cochlear nerve and/or on inner hair cells. Considering that lidocaine alters OHC current (in isolated hair cells) and that lidocaine does not affect endocochlear potential [Laurikainen et al. Acta Otolaryngol (Stockh) 1991: 112: 800-9], the observed CM changes are most likely due to an in vivo effect on OHCs. Thus, the early effect of lidocaine on the cochlea appears to be due to a significant change in organ of Corti function, rather than to direct anesthesia of the cochlear nerve. Later, an independent effect of the drug may occur on neural tissues in the inner ear.

Animals↗

PubMed1997

Isolation of the mouse cDNA homologous to the human EXT1 gene responsible for Hereditary Multiple Exostoses.

We have cloned and sequenced the mouse cDNA homologous to the human Hereditary Multiple Exostoses (EXT1) gene. The mouse homolog shows 94% similarity at the nucleotide level and 99% similarity at the protein level compared to the human gene. The 5' UTRs are unusually conserved for non-coding sequences showing 94% similarity compared to 80% for the 3' UTRs. The high level of evolutionary conservation between the EXT1 proteins as well as the 5' UTR suggests that each plays an important and related role in both species.

Amino Acid Sequence↗