Biomedical subjects

B J Lee

Publications and source records attributed to B J Lee.

At least 37 records · Page 2Linked to original sources

PubMed2001

Terfenadine-beta-Cyclodextrin inclusion complex with antihistaminic activity enhancement.

Terfenadine, an antihistaminic drug, has relatively low bioavailability after oral administration due to its limited solubility in water. To enhance the antihistaminic activity of terfenadine, the terfenadine-beta-cyclodextrin (1:2) inclusion complex was prepared by the neutralization method. The solubility and dissolution of the inclusion complex were carried out, and its antihistaminic activity was then evaluated and compared with terfenadine powder by the passive subcutaneous anaphylaxis method in rats. The formation constant of the inclusion complex was higher at lower pH, while its formation ratio was 1:2 irrespective of pH. For terfenadine, it improved the solubility 200 times and the dissolution rate 5 times. It gave a low histamine level at 30 min, followed by a sustained low level until 60 min, while terfenadine powder gave a low histamine level at 60 min, suggesting that it had faster and more effective antihistaminic activity than terfenadine powder in rats due to fast dissolution and absorption of terfenadine. It is concluded that this inclusion complex enhanced the antihistaminic activity of terfenadine following the enhanced solubility and dissolution of terfenadine.

Algorithms↗

PubMed2001

Detection of the protein-protein interaction between cyclic AMP receptor protein and RNA polymerase, by (13)C-carbonyl NMR.

Cyclic AMP receptor protein (CRP) plays a key role in the transcription regulation of many prokaryotic genes. Upon the binding of cyclic AMP, CRP is allosterically activated, binds to target DNA sites, and interacts with RNA polymerase. Although the protein-protein interaction between CRP and RNA polymerase is known to be important for the transcription initiation of the target genes, its structural understanding is still lacking, particularly due to the high molecular mass (approximately 120 kDa) of the protein complex. We assigned all of the (13)C-carbonyl resonances of methionine residues in CRP by using the double labeling and the enzyme digestion techniques. The result of (13)C-carbonyl NMR experiment on [(13)C'-Met]-CRP in the presence of both cyclic AMP and RNA polymerase alpha subunit showed that the two proteins interact with each other in solution in the absence of DNA via the region around the residues from Met 157 to Met 163 in CRP. The results also showed the effectiveness of the selective labeling and (13)C-carbonyl NMR spectroscopy in the specific detection of the protein-protein interaction between large molecules.

Allosteric Regulation↗

PubMed2001

Full-thickness surgical excision for the treatment of inflammatory linear verrucous epidermal nevus.

Inflammatory linear verrucous epidermal nevus (ILVEN) is a benign cutaneous hamartoma characterized by intensely erythematous, pruritic, inflammatory papules that occur as linear bands along the lines of Blaschko. Because of its chronic and unremitting symptomatology, patients with ILVEN seek medical treatment for relief of discomfort as well as concerns regarding cosmetic appearance. Reported therapeutic approaches include topical agents, dermabrasion, cryotherapy, laser therapy, and partial-thickness excision. Unfortunately, no one therapy has been successful consistently. Medical management is often unsatisfactory, because improvement tends to be temporary. Surgical modalities have met with better success in relief of symptoms but at the risk of marked scarring and a high rate of recurrence. Furthermore, the occurrence of extensive ILVEN or localization to certain anatomic regions has been considered previously a relative contraindication to excision. The authors report 4 patients with extensive ILVEN treated successfully with full-thickness surgical excision. Our report underscores the effectiveness of this surgical modality for the definitive treatment of ILVEN.

Adult↗

PubMed2001

Borna disease virus phosphoprotein binds a neurite outgrowth factor, amphoterin/HMG-1.

The Borna disease virus (BDV) p24 phosphoprotein is an abundant protein in BDV-infected cultured cells and animal brains. Therefore, there is a possibility that binding of the p24 protein to cellular factor(s) induces functional alterations of infected neural cells in the brain. To identify a cellular protein(s) that interacts with BDV p24 protein, we performed far-Western blotting with extracts from various cell lines. Using recombinant p24 protein as a probe, we detected a 30-kDa protein in all cell lines examined. Binding between the 30-kDa and BDV p24 proteins was also demonstrated using BDV p24 affinity and ion-exchange chromatography columns. Microsequence analysis of the purified 30-kDa protein revealed that its N terminus showed complete homology with rat amphoterin protein, which is a neurite outgrowth factor abundant in the brain during development. Mammalian two-hybrid and immunoprecipitation analyses also confirmed that amphoterin is a specific target for the p24 protein in vivo. Furthermore, we showed that infection by BDV, as well as purified p24 protein in the medium, significantly decreased cell process outgrowth of cells grown on laminin, indicating the functional inhibition of amphoterin by interaction with the p24 protein. Immunohistochemical analysis revealed decreased levels of amphoterin protein at the leading edges of BDV-infected cells. Moreover, the expression of the receptor for advanced glycation end products, of which the extracellular moiety is a receptor for amphoterin, was not significantly activated in BDV-infected cells during the process of extension, suggesting that the secretion of amphoterin from the cell surface is inhibited by the binding of the p24 protein. These results suggested that BDV infection may cause direct damage in the developing brain by inhibiting the function of amphoterin due to binding by the p24 phosphoprotein.

Animals↗

PubMed2001

Selective inhibition of selenocysteine tRNA maturation and selenoprotein synthesis in transgenic mice expressing isopentenyladenosine-deficient selenocysteine tRNA.

Selenocysteine (Sec) tRNA (tRNA([Ser]Sec)) serves as both the site of Sec biosynthesis and the adapter molecule for donation of this amino acid to protein. The consequences on selenoprotein biosynthesis of overexpressing either the wild type or a mutant tRNA([Ser]Sec) lacking the modified base, isopentenyladenosine, in its anticodon loop were examined by introducing multiple copies of the corresponding tRNA([Ser]Sec) genes into the mouse genome. Overexpression of wild-type tRNA([Ser]Sec) did not affect selenoprotein synthesis. In contrast, the levels of numerous selenoproteins decreased in mice expressing isopentenyladenosine-deficient (i(6)A(-)) tRNA([Ser]Sec) in a protein- and tissue-specific manner. Cytosolic glutathione peroxidase and mitochondrial thioredoxin reductase 3 were the most and least affected selenoproteins, while selenoprotein expression was most and least affected in the liver and testes, respectively. The defect in selenoprotein expression occurred at translation, since selenoprotein mRNA levels were largely unaffected. Analysis of the tRNA([Ser]Sec) population showed that expression of i(6)A(-) tRNA([Ser]Sec) altered the distribution of the two major isoforms, whereby the maturation of tRNA([Ser]Sec) by methylation of the nucleoside in the wobble position was repressed. The data suggest that the levels of i(6)A(-) tRNA([Ser]Sec) and wild-type tRNA([Ser]Sec) are regulated independently and that the amount of wild-type tRNA([Ser]Sec) is determined, at least in part, by a feedback mechanism governed by the level of the tRNA([Ser]Sec) population. This study marks the first example of transgenic mice engineered to contain functional tRNA transgenes and suggests that i(6)A(-) tRNA([Ser]Sec) transgenic mice will be useful in assessing the biological roles of selenoproteins.

Animals↗

PubMed2001

Expression of telomerase activity in cholesteatoma otitis media.

Telomerase maintains the length of telomeres in immortal cells and is also often associated with cell proliferation. Cholesteatoma epithelium is characterized by a dysregulation with hyperproliferative growth. The study evaluated the telomerase activity in cholesteatoma and normal retro-auricular skin to discover the relationship between telomerase expression and clinical findings. Twenty-two samples of cholesteatoma and 15 samples of retro-auricular skin were obtained from patients undergoing middle-ear surgery. The telomerase activity was detected by the telomerase repeat amplification protocol (TRAP) assay method. Seventeen of the 22 (77.3 per cent) cholesteatoma cases expressed telomerase activity, whereas none of the 15 retro-auricular normal skin (0 per cent) detected telomerase activity. There was no significant difference between telomerase expressions and clinical findings, including hearing level, duration of disease, and the degree of extension (p>0.05). The high expression of telomerase in cholesteatoma suggests that the activation of telomerase may be related to the proliferative nature of cholesteatoma.

Adolescent↗

PubMed2001

Therapeutic effect of hyaluronic acid on experimental osteoarthrosis of ovine temporomandibular joint.

A symptomatic relief by hyaluronic acid (HA, MW: 3.5 x 10(6)), which is synthesized by Streptococcus spp, was investigated in experimental ovine osteoarthrosis. Bilateral osteoarthrosis (OA) of the temporo-mandibular joints (TMJs) was induced by perforating discs and by scrapping subchondral condylar surface. HA was intra-articularly injected into the left joints of 6 sheep on 7, 10, 14, 17 and 21 days after the operation and physiological saline as the control was injected into the contralateral (right) joints on the same day. Three sheep were killed at I month post-operation (MPO) and the remaining three sheep were killed at 3 MPO. Various responses such as proliferation of fibrous tissue, denudation, erosion, osteophyte formation, subcortical cyst formation and ankylosis were observed radiographically and histopathologically. The treatment of HA ameliorated the degenerative changes and lowered the osteoarthrotic score in the left joints at I MPO (9.96 vs 5.81) and 3 MPO (10.86 vs 5.29) compared to the right joints. These results indicate that a repeated intra-articular injection of HA inhibits the progression of OA in ovine TMJs by inducing the development of articular cartilage and by reducing the proliferation of fibrotic tissue.

Animals↗

PubMed2001

Effect of enrofloxacin-Na against pathogens related to the respiratory and alimentary diseases in suckling and weanling piglets.

A field trial was conducted to evaluate effect of enrofloxacin-Na against pathogens related to the respiratory and alimentary diseases in eighty suckling piglets (6-7 days old) and eighty weanling piglets (5-6 weeks old). Respective twenty of the suckling and weanling piglets were assigned to each of 4 experimental groups; control (non-treated), clinical injection dose (CID), 2x clinical injection dose (2CID). and premix. A 0.05 ml (2.5 mg) of enrofloxacin-Na injection (5% solution, 1 ml) per kg body weight of piglets as CID was injected intramuscularly for 3 days and the clinical signs were observed for 9 days. The premix (150 ppm) of enrofloxacin-Na was administered with feed for 7 days ad libitum and the clinical signs were observed for 13 days. The enrofloxacin-Na-treated piglets showed a higher increase in body weight and a lower feed per gain than the control piglets. In addition, the treatment of enrofloxacin-Na, regardless of the route of administration, decreased the incidence rate of diarrhea in suckling piglets and respiratory symptoms in weanling piglets. The isolation index of E. coli and Cl. perfringens during the treatment periods was also lowered by the enrofloxacin-Na treatment in both suckling and weanling piglets. The antibiotics was also evaluated as safe locally and whole bodily as treated by injection or feeding. These results indicate that the newly developed antibiotics, enrofloxacin-Na, is very useful for the prevention and therapy of swine diseases in the pig industry.

Animals↗

PubMed2001

A case of hereditary angioedema associated with idiopathic hypoparathyroidism.

Hereditary angioedema is a rare autosomal dominant disease characterized by the edema of subcutaneous tissues, respiratory tract and bowel. It is caused by the deficiency of C1 esterase inhibitor. Hereditary angioedema may be associated with autoimmune diseases, such as systemic lupus erythematosus, rheumatoid arthritis, autoimmune thyroiditis and glomerulonephritis. We report a 34-year-old male patient with hereditary angioedema who developed idiopathic hypoparathyroidism. Autoimmunity seems to be an important basis of this association and it might be caused by the immune dysfunction due to decreased level of complements; nevertheless, a casual association could not be excluded. To our knowledge, this is the first report of hereditary angioedema in association with idiopathic hypoparathyroidism in the medical literature.

Adult↗

PubMed2001

Effects of nonylphenol, bisphenol A, and their mixture on the viviparous swordtail fish (Xiphophorus helleri).

A number of fish species have been used for studies on endocrine disrupting chemicals (EDCs). However, despite the widespread use of oviparous fish, relatively little attention has been given to viviparous species. This study investigated the effects of EDCs in a viviparous fish and examined the possible usefulness of the fish as an alternative model for the studies on EDCs. Swordtails (Xiphophorus helleri) were exposed to nonylphenol (NP), bisphenol A (BPA), and their mixture. Both short-term (3-d) and relatively long-term (60-d) exposures were carried out using adult male and 30-d-old juvenile fish, respectively. Following the short-term exposure, both NP and BPA caused vitellogenin mRNA expression. Flow cytometric analysis and terminal deoxynucleotidyl transferase assay on the testes of treated fish indicated reproductive damage. Histopathological analysis found degenerative and necrotic cells in seminiferous tubules following the exposure to 100 ppb NP. The testes with lesions were also associated with highly suppressed spermatogenesis. Following the long-term exposure, both NP and BPA exposures significantly affected the growth of swordtails. In all cases, the results showed that the mixture was always more potent than a single chemical and that swordtail fish can be a useful model for the study of endocrine disruptors.

Air Pollutants, Occupational↗

PubMed2001

Flow cytometric evaluation on the age-dependent changes of testicular DNA contents in rats.

An age-dependent cellular change of DNA contents in the testis of Sprague-Dawley rats was investigated by flow-cytometric method. Testicular cell suspensions at the age of 4, 5, 6, 7, 8, 10, 12, 16 and 26 weeks were prepared and stained with propidium iodide. The relative proportions in the number of mature and immature haploid (1n), diploid (2n), S-phase and tetraploid (4n) cells were calculated. The proportion in the number of mature haploid cells was sharply increased to the age of 10 weeks (about 38%), thereafter increased slightly to the level of 42% at the age of 26 weeks. The proportion of immature haploid cells was dramatically increased to the age of 6 weeks, then maintained at the level of 20 to 30% thereafter. The proportion of diploid cells was 64% at the age of 4 weeks, then decreased gradually through the age of 26 weeks. The proportion of S-phase cells was increased to the age of 4 weeks, then maintained at a plateau level to the age of 26 weeks. The proportion of tetraploid cells were about 26% at the age of 4 weeks, then decreased gradually to the age of 26 weeks. These results suggest that the proportions of testicular cells may depend on the age of the rat and that the flow cytometric method may be useful in the evaluation of the spermatogenic status with regard to accuracy and sensitivity.

Animals↗

PubMed2001

Effect of ethylene glycol monoethyl ether on the spermatogenesis in pubertal and adult rats.

The effects of ethylene glycol monoethyl ether (EGEE) on testicular cell populations in pubertal (5 weeks old) and adult (9 weeks old) male rats were investigated by a flow cytometric method. A total of 50 rats (in number, 25 pubertal and 25 adult rats) was divided into 5 experimental groups including 0 (control), 50, 100, 200, and 400 mg EGEE/kg of body weight. The animals were administered by gavage for 4 weeks. In adult rats, the treatment of EGEE at the dose of 400 mg/kg of body weight decreased significantly the populations of haploid, while it increased those of diploid and tetraploid cells. In pubertal rats, the treatment of EGEE at the dose of 400 mg/kg of body weight caused only minimal changes in the relative percent of testicular cell types. These results suggest that the effects of EGEE on testicular function in pubertal rats appear to be less pronounced than in adult rats.

Animals↗

PubMed2000

Survival and stability of bifidobacteria loaded in alginate poly-l-lysine microparticles.

Bifidobacteria-loaded alginate microparticles were prepared by spraying a mixture of alginate and bifidobacteria culture using an air atomization method. Survival and stability of bifidobacteria loaded in microparticles were then evaluated. Survival of bifidobacteria from alginate poly-l-lysine microparticles was significantly increased when MRS broth or yeast extract was added in simulated intestinal fluid (pH 6.8). The number of bifidobacteria gradually increased for 8 h (10(8) cfu/g) and then reached about 10(9)-10(10) cfu/g when incubated over 12 h in intestinal fluid containing 0.5% yeast extract and 0.05% L-cysteine. The survival of bifidobacteria was highly dependent on the pH of the exposing media. When the bifidobacteria was immobilized with alginate or even poly-l-lysine treatment, the survival of bifidobacteria was highly enhanced in the low pH conditions (ca. > 10(8) vs. < 10(3) cfu/g). The stability of free flowing bifidobacteria-loaded alginate poly-l-lysine microparticles was significantly improved during storage at 4 degrees C in a refrigerator when compared to bifidobacteria cultures. The bifidobacteria-loaded alginate poly-l-lysine microparticles could be applied to various dairy products.

Alginates↗

PubMed2000

Structural understanding of the allosteric conformational change of cyclic AMP receptor protein by cyclic AMP binding.

Cyclic AMP receptor protein (CRP) plays a key role in the regulation of more than 150 genes. CRP is allosterically activated by cyclic AMP and binds to specific DNA sites. A structural understanding of this allosteric conformational change, which is essential for its function, is still lacking because the structure of apo-CRP has not been solved. Therefore, we performed various NMR experiments to obtain apo-CRP structural data. The secondary structure of apo-CRP was determined by analyses of the NOE connectivities, the amide proton exchange rates, and the (1)H-(15)N steady-state NOE values. A combination of the CSI-method and TALOS prediction was also used to supplement the determination of the secondary structure of apo-CRP. This secondary structure of apo-CRP was compared with the known structure of cyclic AMP-bound CRP. The results suggest that the allosteric conformational change of CRP caused by cyclic AMP binding involves subunit realignment and domain rearrangement, resulting in the exposure of helix F onto the surface of the protein. Additionally, the results of the one-dimensional [(13)C]carbonyl NMR experiments show that the conformational change of CRP caused by the binding of cyclic GMP, an analogue of cyclic AMP, is different from that caused by cyclic AMP binding.

Allosteric Regulation↗

PubMed2000

Solution structure and membrane-binding property of the N-terminal tail domain of human annexin I.

The conformational preferences of AnxI(N26), a peptide corresponding to residues 2-26 of human annexin I, were investigated using CD and NMR spectroscopy. CD results showed that AnxI(N26) adopts a mainly alpha-helical conformation in membrane-mimetic environments, TFE/water and SDS micelles, while a predominantly random structure with slight helical propensity in aqueous buffer. The helical region of AnxI(N26) showed a nearly identical conformation between in TFE/water and in SDS micelles, except for the orientation of the Trp-12 side-chain, which was quite different between the two. The N-terminal region of the AnxI(N26) helix showed a typical amphipathic nature, which could be stabilized by the neighboring hydrophobic cluster. The helical stability of the peptide in SDS micelles was increased by addition of calcium ions. These results suggest that the N-terminal tail domain of human annexin I interacts with biological membranes in a partially calcium-dependent manner.

Amino Acid Sequence↗

PubMed2000

Structure-expression relationships of the 15-kDa selenoprotein gene. Possible role of the protein in cancer etiology.

Selenium has been implicated in cancer prevention, but the mechanism and possible involvement of selenoproteins in this process are not understood. To elucidate whether the 15-kDa selenoprotein may play a role in cancer etiology, the complete sequence of the human 15-kDa protein gene was determined, and various characteristics associated with expression of the protein were examined in normal and malignant cells and tissues. The 51-kilobase pair gene for the 15-kDa selenoprotein consisted of five exons and four introns and was localized on chromosome 1p31, a genetic locus commonly mutated or deleted in human cancers. Two stem-loop structures resembling selenocysteine insertion sequence elements were identified in the 3'-untranslated region of the gene, and only one of these was functional. Two alleles in the human 15-kDa protein gene were identified that differed by two single nucleotide polymorphic sites that occurred within the selenocysteine insertion sequence-like structures. These 3'-untranslated region polymorphisms resulted in changes in selenocysteine incorporation into protein and responded differently to selenium supplementation. Human and mouse 15-kDa selenoprotein genes manifested the highest level of expression in prostate, liver, kidney, testis, and brain, and the level of the selenoprotein was reduced substantially in a malignant prostate cell line and in hepatocarcinoma. The expression pattern of the 15-kDa protein in normal and malignant tissues, the occurrence of polymorphisms associated with protein expression, the role of selenium in differential regulation of polymorphisms, and the chromosomal location of the gene may be relevant to a role of this protein in cancer.

3' Untranslated Regions↗

PubMed2000

Identification of alternative splicing and negative splicing activity of a nonsegmented negative-strand RNA virus, Borna disease virus.

Borna disease virus (BDV) is a nonsegmented negative-strand RNA virus that belongs to the Mononegavirales. Unlike other animal viruses of this order, BDV replicates and transcribes in the nucleus of infected cells. Previous studies have shown that BDV uses RNA splicing machinery for its mRNA expression. In the present study, we identified spliced RNAs that use an alternative 3' splice site, SA3, in BDV-infected cell lines as well as infected animal brain cells. Transient transfection analysis of cDNA clones of BDV RNA revealed that although SA3 is a favorable splice site in mammalian cells, utilization of SA3 is negatively regulated in infected cells. This negative splicing activity of the SA3 site is regulated by a putative cis-acting region, the exon splicing suppressor (ESS), within the polymerase exon of BDV. The BDV ESS contains similar motifs to other known ESSs present in viral and cellular genes. Furthermore, our results indicated that a functional polyadenylation signal just upstream of the BDV ESS is also involved in the regulation of alternative splicing of BDV. These observations represent the first documentation of complex RNA splicing in animal RNA viruses and also provide new insight into the mechanism of regulation of alternative splicing in animal viruses.

Alternative Splicing↗

PubMed2000

Identification of the gene encoding BmpB, a 30 kDa outer envelope lipoprotein of Brachyspira (Serpulina) hyodysenteriae, and immunogenicity of recombinant BmpB in mice and pigs.

A gene encoding a 30kDa outer envelope protein of the intestinal spirochaete Brachyspira (Serpulina) hyodysenteriae, was cloned and expressed in Escherichia coli strain XLOLR. Five phagemids containing DNA inserts encoding the protein were established and one clone (pSHA) was sequenced. An 816bp hypothetical open reading frame (ORF) was identified, with a potential ribosome binding site (AGGAG), and putative -10 (TATAAT) and -35 (TTGAAA) promoter regions upstream from the ATG start of the ORF. A 12bp inverted repeat sequence, possibly serving as a transcription terminator, was identified downstream from the TAA stop codon. Analysis of the amino acid sequence identified a 19 residue hydrophobic signal peptide, incorporating a potential signal peptidase cleavage site and membrane lipoprotein lipid attachment site. Further analysis of the amino acid usage of this lipoprotein, designated BmpB, showed its possible outer membrane localisation. Comparison of the gene encoding the lipoprotein, bmpB, with GenBank nucleotide sequences showed that it has homology with the gene (plp3) encoding Plp3, an outer membrane lipoprotein of Pasteurella haemolytica (54% identity in 735bp). Comparison of the deduced amino acid sequence with the SWISS-PROT amino acid database revealed greatest homology with the outer membrane lipoproteins (Plp1, 2, 3) of P. haemolytica (34% identity in 242 aa, 37% identity in 250 aa, and 39% identity in 272 aa, respectively), and lipoproteins (rcsF and lipoprotein-28) of E. coli (40% identity in 267 aa and 36% identity in 263 aa, respectively). Three of the recombinant E. coli clones (pSHA, pSHD, and pSHE) were formalinised and used to immunise mice. A bacterin preparation of one recombinant E. coli clone (pSHA) was used to immunise pigs. Sera from these mice and pigs recognised the 30kDa lipoprotein in outer membrane preparations of B. hyodysenteriae, indicating the immunogenicity of recombinant BmpB. Sera from pigs naturally infected with B. hyodysenteriae also reacted with recombinant BmpB expressed in E. coli.

Amino Acid Sequence↗