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Biomedical subjects

L Ni

Publications and source records attributed to L Ni.

At least 73 records · Page 4Linked to original sources
PubMed1995

Phenotypic manifestations of branchio-oto-renal syndrome.

Branchiootorenal (BOR) syndrome is a variable, autosomal-dominant disorder of the first and second embryonic branchial arches, kidneys, and urinary tract. We describe the phenotype in 45 individuals, highlighting differences and similarities reported in other studies. Characteristic temporal bone findings include cochlear hypoplasia (4/5 of normal size with only 2 turns), dilation of the vestibular aqueduct, bulbous internal auditory canals, deep posterior fossae, and acutely-angled promontories.

Abnormalities, Multiple↗
PubMed1995

Pharmacokinetics and toxicity of the human immunodeficiency virus inhibitor 1-ethoxymethyl-6-phenylselenenyl-5-ethyluracil in rodents.

1-(Ethoxymethyl)-6-(phenylselenenyl)-5-ethyluracil (E-EPSeU) has been shown to exhibit potent and selective activity against human immunodeficiency virus type 1 in vitro. The pharmacokinetics of E-EPSeU were characterized after intravenous administration of 5, 10 and 15 mg/kg to rats. Plasma and urine concentrations of E-EPSeU were determined by HPLC. The plasma protein binding of E-EPSeU averaged 86 +/- 4% and the blood: plasma concentration ratio was unity. E-EPSeU concentrations after the 5 mg/kg dose were too low to reliably characterize the pharmacokinetics. The pharmacokinetics of E-EPSeU were independent of dose over the range of 10-15 mg/kg. Plasma concentrations of E-EPSeU declined in a bi-exponential manner with terminal half-life of 0.45 +/- 0.12 h (mean +/- S.D.). The steady-state volume of distribution was 0.091 +/- 0.031 1/kg, suggesting the compound distributed primarily into blood. The systemic clearance (0.63 +/- 0.13 1/h/kg) was moderate and limited, in part, by protein binding. No parent compound was detected in urine. E-EPSeU-related toxicities were observed at high doses. One rat, out of 5, died 4 h after 15 mg/kg of E-EPSeU was administered and two rats administered 20 and 25 mg/kg died within 1 h. Two mice, out of 5, administered 30 mg/kg/day of E-EPSeU intraperitoneally for 6 days died during the experiment, while significant loss of body weight was observed in the surviving mice. However, body weight of the surviving mice returned to control values within 2 weeks after E-EPSeU treatment was stopped.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Linkage of a gene for dominant non-syndromic deafness to chromosome 19.

Inherited hearing impairment can occur either in the presence of other clinical features (syndromic hearing loss, SHL) or in isolation (non-syndromic hearing loss, NSHL). The latter is more common and is highly heterogeneous. To date, six NSHL loci have been mapped. We report the identification of a seventh locus (DFNA4) on chromosome 19q13 and suggest DM kinase as a possible candidate gene.

Chromosome Mapping↗
PubMed1995

Consanguineous nuclear families used to identify a new locus for recessive non-syndromic hearing loss on 14q.

Hearing impairment is inherited most frequently as an autosomal recessive isolated clinical finding (non-syndromic hearing loss, NSHL). Extreme heterogeneity and phenotypic variability in the audiometric profile preclude pooling of affected families and severely hamper gene mapping by conventional linkage analysis. However, in instances of consanguinity, homozygosity mapping can be used to identify disease loci in small nuclear families. This report demonstrates the power of this technique by identifying a locus for recessive NSHL on 14q (DFNB4).

Chromosome Mapping↗
PubMed1995

An autosomal recessive nonsyndromic form of sensorineural hearing loss maps to 3p-DFNB6.

Autosomal recessive nonsyndromic hearing loss (ARNSHL) is the most common form of congenitally acquired inherited hearing impairment. Although numerous loci are believed to exist, only five have been identified. Using a pooled genomic DNA screening strategy, we have identified a sixth locus, DFNB6, on 3p in the interval bounded by D3S1619 and D3S1766.

Alleles↗
PubMed1994

Pharmacokinetics of 2',3'-dideoxy-5-fluoro-3'-thiacytidine in rats.

Although several drugs have shown clinical anti-human immunodeficiency virus activity, reduced activity with long-term use and toxicity make new agents with high therapeutic indices desirable. Racemic cis-2',3'-dideoxy-5-fluoro-3'-thiacytidine (FTC) is a new synthetic nucleoside analogue that is usually potent against human immunodeficiency virus types 1 and 2 and hepatitis B virus in vitro. The purpose of this study was to characterize the preclinical pharmacokinetics of FTC in rats. Rats were administered 10, 50, and 100 mg of FTC per kg of body weight intravenously. Concentrations of FTC in plasma and urine were determined by HPLC. Pharmacokinetic parameters were generated by area/moment analysis. Plasma FTC concentrations declined rapidly in a biexponential fashion, with a terminal half-life of approximately 2 h. The area under the plasma FTC concentration-time curve increased proportionally with increasing dose, and there were no statistically significant differences in pharmacokinetic parameters among the three doses. Thus, the disposition of FTC was independent of dose over the range of 10-100 mg/kg. Since the disposition of FTC was linear, pharmacokinetic parameters were averaged for the three doses. The average total clearance of FTC was 1.91 +/- 0.32 L/h/kg (mean +/- SD), the average renal clearance was 1.08 +/- 0.26 L/h/kg, and the average nonrenal clearance was 0.83 +/- 0.27 L/h/kg. Approximately 55% of the dose of FTC was recovered as unchanged drug in the urine. The steady-state volume of distribution of FTC averaged 2.17 +/- 0.59 L/kg.

Animals↗
PubMed1994

Neutralization epitope of the varicella-zoster virus gH:gL glycoprotein complex.

Varicella-zoster virus (VZV) glycoprotein gpIII is the homolog of herpes simplex virus gH. Through the use of panels of monoclonal antibodies, VZV gpIII is known to possess a complement-independent neutralization epitope which is conformational in nature. Monoclonal antibody to this same epitope, when added postinfection, inhibits both syncytia formation and egress of virus. The nature of the neutralization epitope was investigated to determine whether its formation was dependent on gpIII alone or required a second VZV glycoprotein. To this end, VZV ORF 37 (gH) and VZV ORF 60 (gL homolog) were cloned into a vaccinia virus-pTM1 expression system. Analyses of the transfected products demonstrated that gpIII alone was not fully glycosylated nor was it transported to the cell surface. When both ORF 37 and ORF 60 were cotransfected, the gpIII product was transported to the cell surface, where it formed a neutralization epitope recognized by a previously characterized monoclonal antibody reagent. In summary, the VZV homologs of the herpes simplex virus gH:gL complex included a M(r) 118,000 product (gpIII or gH) and a M(r) 20,000 product (ORF 60 or gL).

Antibodies, Monoclonal↗
PubMed1994

Interferon-gamma promotes cholinergic differentiation of embryonic septal nuclei and adjacent basal forebrain.

In cultured rat embryonic septal nuclei with adjacent basal forebrain, murine interferon-gamma (IFN gamma) produces a striking increase in choline acetyltransferase (ChAT) activity and mRNA. The effect of IFN gamma on cholinergic differentiation is more potent in E14 cultures than in older cultures. IFN gamma does not cause a change in the affinity of ChAT for choline, nor does it affect cell proliferation. Whereas IFN gamma doubles neuronal cell number, the cholinergic cell number increases more than 7-fold. Ameboid microglia respond to IFN gamma with the translocation of p91 to the nucleus. The action of IFN gamma is not mediated by NGF or bFGF. The enhancement of cholinergic expression that occurs with increased cell density may be partly attributable to an endogenous IFN gamma-like molecule, since antibodies to IFN gamma offset the effects of increased cell density.

Animals↗
PubMed1994

Pharmacokinetics of antiviral polyoxometalates in rats.

Polyoxometalates are soluble mineral compounds formed principally of oxide anions and early transition metal cations. The polyoxometalates K12H2[P2W12O48].24H2O (JM 1591), K10[P2W18Zn4(H2O)2O68].20H2O (JM 1596), and [(CH3)3NH]8[Si2W18Nb6O77] (JM 2820) demonstrate potent antiviral activity against human immunodeficiency virus types 1 and 2, herpes simplex virus, and cytomegalovirus in vitro. The preclinical pharmacokinetics of these three compounds were characterized after single-dose intravenous administration of 50 mg/kg to rats. Plasma, urine, and feces were collected for 168 h, and polyoxometalate concentrations were determined by atomic emission. Serum protein binding was measured by equilibrium dialysis. All three compounds were highly bound to serum proteins in a concentration-dependent manner. Total and unbound concentrations of the three compounds in plasma declined in a triexponential manner with terminal half-lives of 246.0 +/- 127.0, 438.4 +/- 129.4, and 32.2 +/- 5.37 h (mean +/- standard deviation) for JM 1591, JM 1596, and JM 2820, respectively. Systemic clearances based on total concentrations in plasma were low, averaging 0.016 +/- 0.002, 0.015 +/- 0.002, and 0.018 +/- 0.003 liter/h/kg for JM 1591, JM 1596, and JM 2820, respectively. The clearances of unbound compounds from plasma averaged 0.966 +/- 0.136, 0.050 +/- 0.005, and 0.901 +/- 0.165 liter/h/kg for JM 1591, JM 1596, and JM 2820, respectively. For JM 1596, the clearance of unbound compound from the kidneys was lower than the glomerular filtration rate (0.086 liter/h/kg), suggesting this polyoxometalate underwent renal tubular reabsorption. However, JM 1591 and JM 2820 appeared to undergo tubular secretion. The fraction of the dose recovered in urine was 11.5, 46.8, and 10.6% for JM 1591, JM 1596, and JM 2820, respectively. Approximately 5% of the dose of each polyoxometalate was recovered in feces. The steady-state volume of distribution based on total concentrations averaged 1.44 liters/kg for JM 1591, 2.39 liters/kg for JM 1596, and 0.59 liter/kg for JM 2820, indicating moderate to wide distribution throughout the body. All three compounds were detected in various tissues 1 week after single-dose administrations, with the highest levels found in the kidneys and liver. The results of this study indicate that the disposition of polyoxometalates is highly dependent on their molecular structure.

Animals↗
PubMed1993

ATPase activity of TyrR, a transcriptional regulatory protein for sigma 70 RNA polymerase.

The TyrR protein of Escherichia coli is the chief transcriptional regulator of several genes essential for aromatic amino acid biosynthesis and transport. It was established in previous studies that this protein binds ATP, that the TyrR.ATP complex has enhanced affinity for tyrosine, and that the susceptibility of the TyrR protein to hydrolysis by trypsin is altered by ATP. Here we show that the TyrR protein has ATPase activity, which is stimulated by tyrosine. In this respect the TyrR protein resembles the transcriptional activator NtrC. The NtrC protein contains an internal polypeptide segment, 220 amino acid residues in length, with a high degree of identity to the TyrR protein, that contains the presumptive ATPase catalytic center.

Adenosine Triphosphatases↗
PubMed1993

A stationary-phase protein of Escherichia coli that affects the mode of association between the trp repressor protein and operator-bearing DNA.

Highly purified preparations of trp repressor (TrpR) protein derived from Escherichia coli strains that were engineered to overexpress this material were found to contain another protein, of 21 kDa. The second protein, designated WrbA [for tryptophan (W) repressor-binding protein] remained associated with its namesake through several sequential protein fractionation steps. The N-terminal amino acid sequence of the WrbA protein guided the design of two degenerate oligonucleotides that were used as probes in the cloning of the wrbA gene (198 codons). The WrbA protein, in purified form, was found by several criteria to enhance the formation and/or stability of noncovalent complexes between TrpR holorepressor and its primary operator targets. The formation of an operator-holorepressor-WrbA ternary complex was demonstrated by gel mobility-shift analysis. The WrbA protein alone does not interact with the trp operator. During the stationary phase, cells deficient in the WrbA protein were less efficient than wild type in their ability to repress the trp promoter. It is proposed that the WrbA protein functions as an accessory element in blocking TrpR-specific transcriptional processes that might be physiologically disadvantageous in the stationary phase of the bacterial life cycle.

Amino Acid Sequence↗
PubMed1993

The immunochemistry of sandwich ELISAs--VI. Greater than 90% of monoclonal and 75% of polyclonal anti-fluorescyl capture antibodies (CAbs) are denatured by passive adsorption.

Quantitative data are presented showing that the method most commonly used to immobilize antibodies in microtiter immunoassays functionally inactivates most of the antibodies. These results were collected using five affinity purified polyclonal antibodies (pAbs) and six monoclonal antibodies (mAbs) specific for fluorescein (FLU) as capture antibodies (CAbs). These CAbs were tested for their ability to capture FLU4.2-BSA after immobilization by passive adsorption, the Protein-Avidin-Biotin-Capture (PABC) system or using previously adsorbed anti-globulins. Results indicate that under optimal conditions, < 10% of monoclonal capture antibody equivalents (CAbeqv) and congruent to 22% of polyclonal CAbeqv remain functional after passive adsorption. Immobilization via the PABC system improved the performance of mAbs by more than five-fold but had less than a two-fold effect on pAbs. Many CAbs immobilized using an anti-globulin retained full activity including the ability to bind two molecules of FLU4.2-BSA/molecule of CAb. The latter result is not necessarily a recommendation for the use of anti-globulin immobilization, since the number of functional CAbeqv per well is not significantly greater than that which can be achieved using passive adsorption.

Adsorption↗
PubMed1992

The physical and functional behavior of capture antibodies adsorbed on polystyrene.

Six monoclonal and two polyclonal antibodies to fluorescein (FLU) were affinity purified and immobilized on Immulon 2 polystyrene as capture antibodies (CAbs): (a) by passive adsorption at pH 9.6, (b) via a streptavidin bridge to a biotinylated carrier molecule, and (c) via an antiglobulin which had been previously adsorbed passively to the polystyrene. Data show that less than 3.0% of the binding sites of monoclonal CAbs and approximately 5-10% of those of polyclonal CAbs were capable of capturing antigen (FLU4.2-BSA) after passive adsorption. Immobilization of CAbs via an antiglobulin or a streptavidin bridge, resulted in the preservation of antibody binding sites to greater than 70% for some monoclonals although immobilization via the streptavidin bridge resulted in the highest number of functional sites/well. The data presented are consistent with studies on other adsorbed proteins which demonstrate that passive adsorption on polystyrene results in the loss of protein function. Furthermore, these data show that generally less than half of the binding sites of antibodies available in solution are available after solid-phase immobilization even when non-adsorptive methods are employed. Some polyclonal anti-FLU also have lower average avidity following passive adsorption compared with CAbs immobilization via a streptavidin bridge. Immunochemical studies revealed that adsorbed polyclonal-CAbs performed like monoclonals when tested with multivalent antigens (FLU10-IgA) but in an expected heterogeneous manner in Scatchard plots when tested using univalent FLU-insulin. This observation implied cross-linking of immobilized CAbs by the multivalent antigen. Because only 5-10% of adsorbed polyclonal CAbs are active, the survivors must be non-randomly distributed in clusters to explain the cross-linking. This was confirmed by scanning electron microscopy which gave rise to the hypothesis that antibodies which retain activity after adsorption, are those present in clusters, i.e., the functional adsorbed CAb is an antibody cluster. Data presented in this report on the behavior of adsorbed CAbs, and reviewed from the work of others for various adsorbed proteins, indicate that the method of passive adsorption at pH 9.6, which is widely used in popular microtiter ELISAs, and which has in many ways revolutionized immunoassay, is a method of protein denaturation. Assayists that utilize passive adsorption of proteins on hydrophobic supports as part of their research need to be cognizant of this phenomenon, while inventors of immunoassay should develop alternative methods of immobilization which do not destroy 90% of the functional activity of solid-phase reactant.

Adsorption↗
PubMed1991

De novo purine nucleotide biosynthesis: cloning, sequencing and expression of a chicken PurH cDNA encoding 5-aminoimidazole-4-carboxamide-ribonucleotide transformylase-IMP cyclohydrolase.

The purH cDNA, encoding 5-aminoimidazole-4-carboxamide-ribonucleotide (AICAR) transformylase-inosine monophosphate cyclohydrolase (ATIC), was cloned by functional complementation of an Escherichia coli purH mutant using a chicken liver cDNA expression library. This represents the first report of the cloning of any eukaryotic ATIC-encoding cDNA (PurH). The avian ATIC mRNA is 2.3 kb long and encodes a protein with an Mr of 64,422. The deduced amino acid sequence is 36% identical to the bacterial purH-encoded enzymes from Bacillus subtilis and E. coli. The avian cDNA was expressed as a glutathione S-transferase (GST) fusion protein that was purified in a single step by affinity chromatography. A novel vector was employed which permits rapid and highly efficient cleavage of the GST fusion protein yielding 10 mg of purified PurH product per liter of bacterial culture. Km values were determined with the purified fusion protein utilizing AICAR and (6-R)N10-formyl-tetrahydrofolate as substrates. These values compare favorably with the isolated avian enzyme, supporting the idea that kinetic, as well as other physical properties of the recombinant fusion protein are similar to the native avian enzyme. Large quantities of purified enzyme and the ability to generate site-directed mutations should make mechanistic studies possible. The recombinant enzyme also affords a simple and reliable approach to identifying new antifolates.

Acyltransferases↗
PubMed1989

Ontogeny of substance P-containing neurons in relation to serotonin-containing neurons in the central nervous system of the mouse.

In order to begin to investigate the developmental factors influencing co-localized neurotransmitters in the central nervous system, we have studied the ontogenesis of coexisting transmitters, substance P and serotonin, in cells of the medullary raphe of the fetal mouse using double-label immunocytochemistry. The results indicate that initial detection of these neurotransmitters occurs at embryonic day 12 in non-overlapping cell populations. Substance P-like immunoreactivity is first co-localized with serotonin in cells of the caudal medullary raphe (raphe pallidus and raphe obscurus) at embryonic day 13. The percentage of serotonin cells containing substance P was estimated at embryonic day 13 and subsequent gestational ages both by double-label immunofluorescence using fluorescein- and rhodamine-conjugated secondary antibodies and also by the elution technique. Double-label immunofluorescence routinely and reliably yielded the highest proportion of co-localized cells. At embryonic day 13, 89% of serotonin cells contained substance P-like immunoreactivity. A consistently high degree of co-localization occurred throughout embryonic development (87% at postnatal day 3) and into adulthood (87%). Colchicine treatment was required at older ages to elicit these data. Unexpectedly, we found that neurons containing substance P-like immunoreactivity in the piriform cortex and in the hypothalamus transiently expressed serotonin immunoreactivity during normal ontogenesis.

Aging↗