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Biomedical subjects

Y Kong

Publications and source records attributed to Y Kong.

At least 109 records · Page 6Linked to original sources
PubMed1997

[Separation and purefied of calf osteocalcin, set up radioimmunoassay for osteocalcin].

OBJECTIVE AND METHODS: We extracted and purified osteocalcin (BGP) from calf femur by gel filtration, FPLC and HPLC subsequently. The rabbits were immunized by this antigen and produced antibody. With this antibody (final 1: 35,000 dilution) we established RIA for BGP. This assay is stable and sensitive, the kd was 2.29 x 10(11). The intra assay variation was 2.1% and the inter assay variaton was 5.6%. It has no crossreaction with insulin, parathyroid hormone or calcitonin. RESULTS: The serum BGP values were detected in normal subjects and six kinds of bone metabolic diseases. In 81 normal subjects, the average serum BGP level was (5.3 +/- 1.5) ng/ml and in postmenopausal osteoporosis the serum BGP level [(6.2 +/- 1.9) ng/ml, n = 42] was higher than the normal (P < 0.05). The serum BGP levels were increased much in primary hyperparathyroidism [(12.8 +/- 7.0) ng/ml, n = 21, P < 0.001]. Increased values of BGP were also found in patients with chronic renal failure [(8.5 +/- 2.4) ng/ml, n = 18, P < 0.01] and Paget's disease [(6.7 +/- 2.2) ng/ml, n = 12, P < 0.05]. Decreased serum BGP values were found in Cushing syndrome [(2.9 +/- 0.9) ng/ml, n = 16, P < 0.001] and hypoparathyroidism [(3.4 +/- 1.1) ng/ml, n = 42, P < 0.001]. This result showed that in diseases with high bone turnover, the serum BGP levels were increased and in diseases with low bone turnover, the serum BGP levels were decreased. CONCLUSION: The data suggest that serum BGP is a sensitive marker for bone turnover. It is very improtant in studying bone metabolic diseases.

Adult↗
PubMed1997

Cysteine protease activities during maturation stages of Paragonimus westermani.

In mature Paragonimus westermani, specific activity of parasitic cysteine protease declines. To clarify which of the known 17-, 27-, and 28-kDa enzyme activities is decreased, the cysteine proteases were purified from the crude extracts of metacercariae, 4- and 7-wk juveniles, and 16-wk adults by gel filtration, ion-exchange, and affinity matrix chromatographies; the enzyme activity was monitored with the fluorogenic substrate, Cbz-phe-arg-AMC. In addition to 3 known enzymes, 2 other cysteine proteases at 15 and 53 kDa were identified in juveniles and adults and were purified. The 2 novel enzymes were most active in 0.1 M ionic strength and pH 5-6 and were inhibited by N-(N-[L-3-transcarboxyrane-2-carbonyl]-L-leucyl)agamatine, iodoacetamide, and leupeptin. Of the 5 enzymes, specific activities of metacercarial 27- and 28-kDa enzymes were lowered from metacercaria to 16 wk. Between 4 and 16 wk, activities of 3 cysteine proteases of juveniles and adults were additionally exhibited. The activity changes of 5 different cysteine proteases may be associated with migration and immune evasion during the maturation stage of P. westermani when the parasite environment is changing.

Animals↗
PubMed1997

Epidemiology of paragonimiasis in Korea.

In Korea, soybean-sauce soaked freshwater crabs (kejang) have been a favorite delicacy when eating a bowl of rice. This traditional food has been a main source of human paragonimiasis. Until the late 1960s, human paragonimiasis had been prevalent; at least two million people had contracted the infection as determined by intradermal tests. About 40% of these were egg positive. In the turmoil of the green revolution and industrialization in the 1970s/1980s, ecological damage occurred widely. In many streams, populations of snail and crustacean hosts were reduced to levels almost of extinction. Population reduction of the intermediate hosts was followed by lowered endemicity. Attitudes of people, changed during the period, also reduced chances of paragonimiasis. Survey data in the 1990s indicated that prevalence of human paragonimiasis has lowered to about one 100th of that in the early 1970s. In a referral system, however, about one hundred clinical cases have been diagnosed annually by antibody test (ELISA), undertaken for clinical differentiation from tuberculosis. At least 10% of freshwater crabs sold in local markets are infected with the metacercariae. Paragonimiasis control has benefited in Korea mainly by the untoward effects of water pollution. To place P. westermani infections as enzootic, health education and surveillance systems should have a priority.

Animals↗
PubMed1996

Theory of multivalent binding in one and two-dimensional lattices.

Ligand binding to a linear lattice composed of N sites, under general conditions of cooperativity and number of sites covered upon binding, m, is approached in terms of the theory of contracted partition functions. The partition function of the system obeys a recursion relation leading to a generating function that provides an exact analytical solution for any case of interest. Site-specific properties of the lattice are derived from simple transformations of the analytical expressions. The McGhee-von Hippel model is obtained as a special case in the limit N --> infinity. The derivation is straightforward and involves no combinatorial arguments. Partition functions and site-specific properties are also derived for the case of non-cooperative binding to a two-dimensional torus of length N, containing s sites in its section for a total of sN sites. The torus provides a relevant model for ligand binding to double-stranded DNA (s = 2) or protein helices (s = 3,4). It is proved that non-cooperative binding to the two-dimensional torus can mimic cooperative binding to a one-dimensional linear lattice when m = s. The dimensional embedding of the lattice and the geometry of interaction of its sites play a crucial role in defining the binding properties of the system accessible to experimental measurements. Hence, caution must be exercised in the interpretation of Scatchard plots in terms of the one-dimensional McGhee-von Hippel model, especially when m < or = 4 and the geometry of the system is clearly two-dimensional.

Journal Article↗
PubMed1996

An oligosaccharide sialyl-Lewis(x) analogue does not reduce myocardial infarct size after ischemia and reperfusion in dogs.

BACKGROUND: Polymorphonuclear leukocytes, particularly neutrophils, are important mediators of ischemia/reperfusion-induced myocardial and coronary vascular injury. The selectin family of glycoprotein receptors mediates neutrophil "rolling," a loose, transient adhesion to the coronary endothelium that precedes the firmer adhesion associated with cardiovascular injury. The oligosaccharide sialyl-Lewis(x) (SLe(x)) is the probable neutrophil counterligand for endothelial E- and P-selectin. Administration of analogues of SLe(x) could potentially prevent neutrophil rolling by competing for the selectin-adhesion sites. We investigated the effects of treatment with an analogue of SLe(x) in a chronic canine model of ischemia/reperfusion. METHODS AND RESULTS: Anesthetized mongrel dogs were subjected to 90 minutes of ischemia through occlusion of the left anterior descending coronary artery and 48 hours of reperfusion. Five minutes before the onset of reperfusion, dogs received either the SLe(x) analogue CY-1503 at a dose of 20 mg/kg or normal saline. Myocardial infarct size was measured through triphenyltetrazolium chloride staining, and polymorphonuclear leukocyte accumulation was evaluated through measurement of cardiac myeloperoxidase activity. After adjustment for blood flow, the mean infarct size of control dogs (44.7 +/- 4.2%) was not different from that of treated dogs (33.4 +/- 4.0%, P = .06), although there was a trend toward a slightly lower value in the treated dogs. Myeloperoxidase activity was not different in the infarcted myocardium of the treated group compared with that of the control group (2.7 +/- 0.71 treated versus 1.08 +/- 0.41 units/mg protein control, P = .06). CONCLUSIONS: We conclude that CY-1503 does not substantially or consistently reduce myocardial infarct size or neutrophil accumulation in dogs subjected to ischemia followed by a prolonged period (48 hours) of reperfusion.

Animals↗
PubMed1996

Nucleotide sequences of MHC class I introns 1, 2, and 3 in humans and intron 2 in nonhuman primates.

HLA-class I genes are the most polymorphic genetic system yet known. The polymorphic substitutions are mostly located in exon 2 and 3, encoding alpha 1 and alpha 2 domains, respectively, which are involved in peptide binding and T cell receptor interaction. In this study, we present the sequences of the introns neighboring the polymorphic exons in humans with few examples from nonhuman primates. In general, intron sequences are found to be less polymorphic than the adjacent exons, displaying numerous locus-specific and group-specific sites. These sequences will provide important information for developing DNA based typing strategies for HLA-class I alleles.

Animals↗
PubMed1996

Stable expression of anti-HPV 16 E7-ribozyme in CV-1 cell lines.

The HPV16 (human papilloma virus type 16) E7 gene product, an oncoprotein, has been considered to be involved in the pathogenesis of anogenital cancer, particularly of cervical cancer. In order to evaluate the effect of suppression of the expression of the E7 gene in CV-1 cells by ribozyme, Rz523 with a transacting ribozyme targeted to the E7 RNA and two processing ribozyme genes at the 5' and 3' flank was cloned into the eukaryotic expression plasmid pREP9 under the control of RSV-LTR promoter. The resultant plasmid pRSV-Rz523 was transfected into CV-1 cells by calcium phosphate coprecipitation. The expression of the ribozyme in G418-resistant cells was detected by dot-blot hybridization. Ribozymes stably expressed in the CV-1 cells were at a level of 9.0 pmol per 10(6) cells, in which the active ribozyme molecules were more than 50 fmol per 10(6) cells. The result of RNase protection assay showed that the steady-state level of the E7 RNA fragment in CV-1 cell lines was significantly reduced by about 90% in ribozyme-expressing cells. In contrast, the antisense control plasmid pRSV-AE7 only exhibited about 20%. This result implicated the possibility of reversing the malignant phenotype of cervical cancer by means of suppressing the expression of the E7 gene with ribozyme.

Cell Line↗
PubMed1995

HNF3 binds and activates the second enhancer, ENII, of hepatitis B virus.

The basic functional unit of hepatitus B virus (HBV) enhancer II (ENII) is located within nt 1687-1774, which is defined as the B fragment in our previous papers. A major trans-acting factor binding site has been identified within the B fragment. The sequence corresponding to this binding site was named B2. In this paper, several point mutations were introduced into the B2 subunit by PCR-mediated site-directed mutagenesis. CAT analysis indicated that the TGTTTGTTT motif within the B2 subunit was critical for the activity of ENII. Mutations of individual nucleotides within this motif could decrease the activity of ENII. Electrophoresis mobility shift assay revealed that the liver-enriched transcription factors hepatocyte nuclear factor (HNF) 3 alpha and HNF3 beta bound to the B2 subunit specifically and the TGTTTGTTT motif was essential for DNA-protein interaction. Anti-HNF3 alpha and anti-HNF3 beta antisera could block such binding ability. Moreover, HNF3 beta could switch on the activity of ENII in HeLa cells and the activity of ENII could be suppressed by antisense HNF3 alpha and antisense HNF3 beta mRNA in HepG2 cells. These results prompted the conclusion that HNF3 was crucial for the liver-specific activity of ENII, which in turn contributed significantly to the liver specificity of HBV.

Base Sequence↗
PubMed1995

Locus-specific amplification of HLA class I genes from genomic DNA: locus-specific sequences in the first and third introns of HLA-A, -B, and -C alleles.

We have identified locus-specific sequences in the first and third introns flanking the polymorphic second and third exons of HLA class I genes. PCR primers derived from these conserved sequences produced DNA fragments of the expected sizes for each of the HLA-A, -B, and -C loci in the amplification of genomic DNA. PCR products generated using each of the locus-specific sets of primers displayed exquisite locus specificity, as assessed by hybridization with oligonucleotide probes specific for ten classical and non-classical HLA class I genes. Amplification with these primer sets was effective and specific for the HLA alleles tested under the given PCR conditions. When hybridized with oligonucleotides derived from shared polymorphic sequence motifs, reaction patterns of PCR products from each locus were precisely as expected from published or database sequences. Chemiluminescent signals generated from digoxygenin-ddUTP-labeled probes were even for all samples and as strong as those obtained in MHC class II typing. These locus-specific primer sets derived from intron sequences provide an effective means to amplify genomic DNA which will facilitate PCR-based HLA class I typing methods. This will also allow HLA class I typing to be conducted with greater precision, at lower cost, and faster than previously described class I typing methodologies.

Alleles↗
PubMed1995

Ras p21Val inhibits myogenesis without altering the DNA binding or transcriptional activities of the myogenic basic helix-loop-helix factors.

MRF4, MyoD, myogenin, and Myf-5 are muscle-specific basic helix-loop-helix transcription factors that share the ability to activate the expression of skeletal muscle genes such as those encoding alpha-actin, myosin heavy chain, and the acetylcholine receptor subunits. The muscle regulatory factors (MRFs) also exhibit the unique capacity to initiate the myogenic program when ectopically expressed in a variety of nonmuscle cell types, most notably C3H10T1/2 fibroblasts (10T1/2 cells). The commitment of myoblasts to terminal differentiation, although positively regulated by the MRFs, also is controlled negatively by a variety of agents, including several growth factors and oncoproteins such as fibroblast growth factor (FGF-2), transforming growth factor beta 1 (TGF-beta 1), and Ras p21Val. The molecular mechanisms by which these varied agents alter myogenic terminal differentiation events remain unclear. In an effort to establish whether Ras p21Val represses MRF activity by directly targeting the MRF proteins, we examined the DNA binding and transcription activation potentials of MRF4 and MyoD when expressed in 10T1/2 cells or in 10T1/2 cells expressing Ras p21Val. Our results demonstrate that Ras p21Val inhibits terminal differentiation events by targeting the basic domain of the MRFs, and yet the mechanism underlying this inhibition does not involve altering the DNA binding or the inherent transcriptional activity of these regulatory factors. In contrast, FGF-2 and TGF-beta 1 block terminal differentiation by repressing the transcriptional activity of the MRFs. We conclude that the Ras p21Val block in differentiation operates via an intracellular signaling pathway that is distinct from the FGF-2 and TGF-beta 1 pathways.

Cell Differentiation↗
PubMed1995

A cysteine protease of Paragonimus westermani eggs.

Protease activity was identified in crude extracts of Paragonimus westermani eggs which were purified from infected dog lungs, isolated on 14 weeks after metacercarial challenge. The eggs were used after removing possibly contaminated host or worm tissues on their shell surfaces. In the crude egg extracts, high proteolytic activities against carboxybenzoyl-phenylalanyl-arginyl-4-methoxy-beta-naphthylamide (Cbz-phe-arg-MNA) and Azocoll were detected whereas those against succinyl-alanyl-prolyl- phenylalanyl-p-nitroanilide (Suc-ala-pro-phe-pNA) were not revealed. The enzyme exhibited the maximal activity at pH 6. Its activity was inhibited by specific cysteine protease inhibitors, 10(-5) M 1-trans-epoxysuccinylleucylamido (4-guanidino) butane (E-64) and 1 mM iodoacetamide (IAA) while potentiated by 6.5-fold in the presence of 2.5 mM dithiothreitol (DTT). When the enzyme was purified partially by Sephacryl S-300 High Resolution gel filtration, it migrated as a single homogeneous band at 35 kDa. The 35 kDa cysteine protease has been recognized neither in the metacercariae nor in the adult. These findings indicated the presence of at least one protease of cathepsin family in immature eggs of P. westermani.

Animals↗
PubMed1995

Molecular cloning and expression of a human secretin receptor.

Secretin is a 27-amino acid neuroendocrine peptide that stimulates fluid and electrolyte secretion in the gastrointestinal tract, activates tyrosine hydroxylase activity in the central nervous system, and affects cardiac and renal function. Specific receptors for secretin have been previously characterized on neuroblastoma cells, pancreatic acini, gastric glands, and liver cholangiocytes. We report here the isolation of a 1616-base pair cDNA from human lung tissue that encodes a 440-amino acid, 50-kDa, G protein-coupled human secretin receptor (HSR), with homology of 80% with the rat secretin receptor and 37% with the human type I vasoactive intestinal peptide receptor. Northern blot analysis of human tissue mRNA revealed that the relative intensity for expression of a 2.1-kilobase HSR transcript was pancreas > kidney > small intestine > lung > liver, with trace levels in brain, heart, and ovary. Stable transfectants of HSR in human embryonic kidney 293 cells, termed 293S12, expressed 10(5) binding sites/cell for 125I-secretin, with an apparent Kd of 3.2 nM. Vasoactive intestinal peptide, pituitary adenylyl cyclase-activating peptide-38, and glucagon were less potent (by 3 orders of magnitude) than secretin in competitively inhibiting 125I-secretin binding to 293S12 cells. Secretin evoked concurrent dose-dependent increases in intracellular cAMP and calcium levels in 293S12 cells and stimulated a 4-fold increase in phosphatidylinositol hydrolysis. Thus, the HSR expressed by stable transfectants can couple to two distinct intracellular signaling pathways.

Amino Acid Sequence↗
PubMed1995

Excystment of Paragonimus westermani metacercariae by endogenous cysteine protease.

To infect definitive or paratenic hosts, metacercariae of Paragonimus westermani should excyst in the host intestine. Optimum conditions for the excystment have been known to be pH 8-9 and a temperature of 40 C. Under these conditions, excystment of P. westermani metacercariae was accelerated in the presence of 1 mM dithiothreitol (DTT). The DTT acceleration was antagonized dose-dependently by cysteine protease inhibitors of L-trans-epoxysuccinylleucylamido(4-guanidino)butane (E-64, 2-20 microM) or leupeptin (0.1-1 mM), suggesting that certain cysteine proteases of the metacercaria are involved in excystment. Protease activities were detected in excretory-secretory products (ESP) of newly excysted metacercariae. Two distinct proteases were purified by DEAE anion-exchange chromatography of the ESP. While a 27-kDa protease exhibited endodipeptidolytic activity at pH 5-8.5 and remained stable at neutral pH for 3 days, the 28-kDa enzyme was stable at pH 5-7.5, with lower activity at pH 8.5. Both proteases hydrolyzed collagen, fibronectin, and myosin within 1 hr at pH 8. These results suggest that cysteine proteases secreted by P. westermani metacercariae modulate excystment.

Animals↗
PubMed1995

HBV surface antigen proteins with deletions in the preS region.

Four pairs of HBV surface antigen genes, in which the preS region was partially deleted, were constructed by the polymerase chain reaction (PCR). The comparison of the levels of the expression in mammalian cells of these genes and the ones constructed before, and the properties of these gene products showed that the missing of a part of the preS region did not affect the overall spatial structure of the S region and the surface localization of the preS region. The removal of the preS1 retention sequence (a. a. 2-19) alleviated significantly the shelter of the major antigenic determinants in the S region by the preS sequence. It was found that the long preS region seriously impaired the secretion of the surface antigen proteins from mammalian cells. In addition to the previously reported preS1 retention sequence, the preS1 sequence (a.a. 48-65) may also inhibit the secretion of the surface antigen proteins, whereas the preS2 region exerts no major influence on the retention of the large surface antigen protein. One of the expressed surface antigen proteins, in which the preS1 sequence (a.a. 21-47) and the S region were directly fused, deserves further study and may be developed into a new HBV vaccine which contains the preS1 binding site for hepatocyte receptors due to its stability, fine secretability and strong preS1 antigenicity.

Animals↗
PubMed1995

Study on nucleic acid synthesis in mouse retina and retinal pigment epithelium by radioautography.

The ocular tissues of ddY mice from embryonic 9 to postnatal day 14 were labelled in vitro with 3H-thymidine or 3H-uridine. After radioautography, the radioautographs were observed by light and electron microscopes. The labeling indices (L.I.) with 3H-thymidine or grain numbers with 3H-uridine were calculated and the area (microns 2) of cell organelles in both labelled and unlabelled retinal cell with 3H-thymidine were measured by image analysis. The L.I. of retinal cells were low in the beginning then increased and reached a peak at P3, while the L.I. of retinal pigment epithelium (RPE) increased and reached a peak at P7. The L.I. of both retinal and PE cells disappeared at P14. However, the L.I. of anterior region were more than those of equaterial and posterior regions in both retinal and PE cells. The results of the image analysis showed that the average area of cell organelles, both labelled and unlabelled retinal cells decreased from E9 to P14. The grain numbers of retina labelled with 3H-uridine increased until P1 then decreased, while the numbers of RPE increased and reached a peak at P3 whereafter they declined. No significant differences were detected among the three regions.

Animals↗
PubMed1995

[Two case of androgen-secreting ovary tumor].

Two cases of androgen-secreting ovary tumor were reported. patients' chief complaints were menoxemia and hirsutism; clitorism was found by physical examination. Their serum testosterone (T) were high (10.2-6.7nmol/L) and could not be suppressed to normal range by middle-dose dexamethasone inhibiting test. ACTH stimulating test could not stimulate the high serum T to raise futher, but HCG stimulating test could increase serum T to higher level. Pelvic examination, type B ultrasonic and CT scan confirmed a mass on one of the ovaries. Sertoli-Leydig cell tumor and gynandroblastoma were proved respectively by pathology. After removing the tumor, their serum T were returned to normal level.

Adult↗