Biomedical subjects

H Song

Publications and source records attributed to H Song.

At least 37 records · Page 2Linked to original sources

PubMed2001

Global gene expression analysis to identify molecular markers of uterine receptivity and embryo implantation.

Infertility and spontaneous pregnancy losses are an enduring problem to women's health. The establishment of pregnancy depends on successful implantation, where a complex series of interactions occurs between the heterogeneous cell types of the uterus and blastocyst. Although a number of genes are implicated in embryo-uterine interactions during implantation, genetic evidence suggests that only a small number of them are critical to this process. To obtain a global view and identify novel pathways of implantation, we used a dual screening strategy to analyze the expression of nearly 10,000 mouse genes by microarray analysis. Comparison of implantation and interimplantation sites by a conservative statistical approach revealed 36 up-regulated genes and 27 down-regulated genes at the implantation site. We also compared the uterine gene expression profile of progesterone-treated, delayed implanting mice to that of mice in which delayed implantation was terminated by estrogen. The results show up-regulation of 128 genes and down-regulation of 101 genes after termination of the delayed implantation. A combined analysis of these experiments showed specific up-regulation of 27 genes both at the implantation site and during uterine activation, representing a broad diversity of molecular functions. In contrast, the majority of genes that were decreased in the combined analysis were related to host immunity or the immune response, suggesting the importance of these genes in regulating the uterine environment for the implanting blastocyst. Collectively, we identified genes with recognized roles in implantation, genes with potential roles in this process, and genes whose functions have yet to be defined in this event. The identification of unique genetic markers for the onset of implantation signifies that genome-wide analysis coupled with functional assays is a promising approach to resolve the molecular pathways required for successful implantation.

Animals↗

PubMed2001

Analysis of TGF-beta type I receptor for mutations and polymorphisms in head and neck cancers.

Transforming growth factor-beta receptor (TbetaR)-dependent signals are critical for cell growth and differentiation and are often disrupted during tumorigenesis. The entire coding region of TbetaR-I and flanking intron sequences from 30 head and neck carcinomas were examined for alterations using "Cold" SSCP and direct sequencing. No somatic point mutations were found in the TbetaR-I gene. In contrast, 14 polymorphic sequence changes were detected in TbetaR-I in 13 (43%) of the samples, including eight (27%) nucleotide alterations identified as polymorphisms in an exon-1 (GCG)(9) microsatellite repeat, a previously reported tumor susceptibility allele. A nine base pair deletion was found in 23% of the samples including five heterozygous and two homozygous deletions as well as single homozygous 12bp deletion. Additionally, six heterozygous polymorphisms in intronic sequences were determined, including one heterozygous C/A genotype at the +82 nucleotide position of the intron-5 intervening sequence (IVS), and five heterozygous G/A genotypes within intron-7 at the +24 nucleotide position. Exon-1 polymorphisms in the (GCG)(9) microsatellite region of the TbetaR-I gene and their association with head/neck cancers, suggest that development of these cancers may be a direct consequence of loss of responsiveness to TGF-beta mediated growth inhibition.

Activin Receptors, Type I↗

PubMed2001

Molecular identification of IgE-dependent histamine-releasing factor as a B cell growth factor.

The culture supernatants of LK1 cells, murine erythroleukemia cells, showed B cell-stimulating activity. Purification and NH(2)-terminal sequence analysis revealed that one of the candidates was murine IgE-dependent histamine-releasing factor (IgE-HRF), which is known to induce histamine from basophils. Recombinant IgE-HRF (rHRF) obtained from Escherichia coli- or 293-transformed embryonal kidney cells was tested for B cell-stimulating activity. Both rHRFs stimulated B cell proliferation in a dose-dependent manner. However, boiling or anti-HRF Ab abolished the B cell stimulatory effects of rHRF. Recombinant HRF showed strong synergistic effects with IL-2, IL-4, and IL-5 for B cell activation, with maximal activity in the presence of anti-CD40 AB: Recombinant HRF increased MHC class II expression of B cells. It also increased Ig production from B cells. Treatment with polymyxin B, a neutralizing peptide antibiotic of LPS, did not reduce the activity of rHRF. In addition, FACS analysis using PE-conjugated rHRF showed that HRF bound to B cells. Recombinant HRF up-regulated the expression of IL-1 and IL-6 in B cells. In vivo administration of rHRF or the cDNA for rHRF increased total and Ag-specific Ig synthesis. Taken together, these results indicate that HRF stimulates B cell activation and function.

Animals↗

PubMed2001

ATTEMPTS: a heparin/protamine-based prodrug approach for delivery of thrombolytic drugs.

The aim of this study is to develop a heparin/protamine-based prodrug system for the controlled delivery of enzyme such as tissue-type plasminogen activator (tPA). This approach, termed antibody targeted, triggered, electrically modified prodrug-type strategy (ATTEMPTS), would permit antibody-directed administration of inactive tPA, and allow a subsequent triggered release of the active tPA at the target site. Cation-modified tPA (mtPA) was attached to a heparin--antifibrin complex via ionic interaction. The active tPA can be subsequently released by the addition of protamine, a competitive heparin inhibitor. Anti-fibrin IgG was conjugated to heparin via an end-point attachment to form the heparin--antifibrin--complex which provides the targeting efficiency of the final heparin--mtPA complex. Cation-modification was performed either by chemical conjugation by linking (Arg)(7)Cys to tPA with N-succinimidy-3-(2-pyridyldithio) propionate or by recombinant DNA method. Results show that the chemical modification process did not significantly alter specific activity of tPA with regard to plasminogen activation, fibrin-binding ability, and response toward fibrinogen. Expressed modified tPA (EmtPA) produced by recombinant DNA methods retained the same catalytic activity of the parent tPA, as well as a dynamic catalytic behavior depending upon the presence of heparin and protamine. Both types of modified tPA, especially the mtPA demonstrated a significantly higher affinity toward heparin or heparin--antifibrin complex than native tPA. In addition, the complexes of mtPA--heparin did not yield any intrinsic clot lysis activity owing to the blockage of the active site of tPA by attached heparin. On the other hand, heparin-induced inhibition of both mtPA and EmtPA activity was reversed by adding protamine, as confirmed by chromogenic and in vitro clot lysis assays. These results suggested that a heparin/protamine-based tPA delivery system may be a useful tool to improve current thrombolytic therapeutic status, by both precisely regulating the release of active tPA and aborting the associated bleeding risk. Alternatively, this ATTEMPTS approach could also be used to deliver enzyme drugs while diminishing their associated toxic effects.

Antifibrinolytic Agents↗

PubMed2001

Immunogenicity of recombinant GA733-2E antigen (CO17-1A, EGP, KS1-4, KSA, Ep-CAM) in gastro-intestinal carcinoma patients.

Targeting the GA733 antigen (also known as CO17-1A, EGP, KS1-4, KSA, Ep-CAM) by monoclonal antibody CO17-1A or anti-idiotypic antibodies mimicking the CO17-1A or GA733 epitope has induced prolonged survival and specific immune responses to the antigen, respectively, in colorectal cancer (CRC) patients. In pre-clinical studies in mice and rabbits, recombinant baculovirus-derived GA733-2E antigen was superior to anti-idiotypic antibodies at modulating specific immune responses. Our aim was to evaluate the immunogenicity and potential toxicity of alum-precipitated GA733-2E in a phase I trial in patients with resected CRC or pancreatic cancer. Six patients with advanced pancreatic carcinoma and 6 with CRC Dukes' stage A, B or C received between 4 and 7 doses of alum-precipitated GA733-2E at 50, 200 or 800 microg/dose at monthly intervals. Antibody binding to GA733-2E or antigen-positive CRC cells was determined, as were antigen-specific proliferative, cytolytic T-lymphocyte and delayed-type hypersensitivity responses. Six of the 12 patients developed antigen-specific humoral immune responses after immunotherapy, and 8 developed cellular immune responses. The overall immune response rate, including patients with humoral and/or cellular immune responses, was 83%. Median overall survival of the CRC and pancreatic cancer patients was 39.8 and 11.2 months, respectively. Following 18 years of single-epitope targeting of the GA733 antigen, immunization of patients against multiple epitopes of the antigen frequently induces an immune response in the absence of significant toxicity, despite relatively widespread expression of this antigen on normal epithelial cells.

Aged↗

PubMed2001

Dysregulated cannabinoid signaling disrupts uterine receptivity for embryo implantation.

The mechanisms by which synchronized embryonic development to the blastocyst stage, preparation of the uterus for the receptive state, and reciprocal embryo-uterine interactions for implantation are coordinated are still unclear. We show in this study that preimplantation embryo development became asynchronous in mice that are deficient in brain-type (CB1) and/or spleen-type (CB2) cannabinoid receptor genes. Furthermore, whereas the levels of uterine anandamide (endocannabinoid) and blastocyst CB1 are coordinately down-regulated with the onset of uterine receptivity and blastocyst activation prior to implantation, these levels remained high in the nonreceptive uterus and in dormant blastocysts during delayed implantation and in pregnant, leukemia inhibitory factor (LIF)-deficient mice with implantation failure. These results suggest that a tight regulation of endocannabinoid signaling is important for synchronizing embryo development with uterine receptivity for implantation. Indeed this is consistent with our finding that while an experimentally induced, sustained level of an exogenously administered, natural cannabinoid inhibited implantation in wild-type mice, it failed to do so in CB1(-/-)/CB2(-/-) double mutant mice. The present study is clinically important because of the widely debated medicinal use of cannabinoids and their reported adverse effects on pregnancy.

Amino Acid Sequence↗

PubMed2001

Somatic INK4a-ARF locus mutations: a significant mechanism of gene inactivation in squamous cell carcinomas of the head and neck.

The INK4a-ARF locus is located on human chromosome 9p21 and is known to encode two functionally distinct tumor-suppressor genes. The p16(INK4a) (p16) tumor-suppressor gene product is a negative regulator of cyclin-dependent kinases 4 and 6, which in turn positively regulate progression of mammalian cells through the cell cycle. The p14(ARF) tumor-suppressor gene product specifically interacts with human double minute 2, leading to the subsequent stabilization of p53 and G(1) arrest. Previous investigations analyzing the p16 gene in squamous cell carcinomas of the head and neck (SCCHNs) have suggested the predominate inactivating events to be homozygous gene deletions and hypermethylation of the p16 promoter. Somatic mutational inactivation of p16 has been reported to be low (0-10%, with a combined incidence of 25 of 279, or 9%) and to play only a minor role in the development of SCCHN. The present study examined whether this particular mechanism of INK4a/ARF inactivation, specifically somatic mutation, has been underestimated in SCCHN by determining the mutational status of the p16 and p14(ARF) genes in 100 primary SCCHNs with the use of polymerase chain reaction technology and a highly sensitive, nonradioactive modification of single-stranded conformational polymorphism (SSCP) analysis termed "cold" SSCP. Exons 1alpha, 1beta, and 2 of INK4a/ARF were amplified using intron-based primers or a combination of intron- and exon-based primers. A total of 27 SCCHNs (27%) exhibited sequence alterations in this locus, 22 (22%) of which were somatic sequence alterations and five (5%) of which were a single polymorphism in codon 148. Of the 22 somatic alterations, 20 (91%) directly or indirectly involved exon 2, and two (9%) were located within exon 1alpha. No mutations were found in exon 1beta. All 22 somatic mutations would be expected to yield altered p16 proteins, but only 15 of them should affect p14(ARF) proteins. Specific somatic alterations included microdeletions or insertions (nine of 22, 41%), a microrearrangement (one of 22, 5%), and single nucleotide substitutions (12 of 22, 56%). In addition, we analyzed the functional characteristics of seven unique mutant p16 proteins identified in this study by assessing their ability to inhibit cyclin-dependent kinase 4 activity. Six of the seven mutant proteins tested exhibited reduced function compared with wild-type p16, ranging from minor decreases of function (twofold to eightfold) in four samples to total loss of function (29- to 38-fold decrease) in two other samples. Overall, somatic mutation of the INK4a/ARF tumor suppressor locus, resulting in functionally deficient p16 and possibly p14(ARF) proteins, seems to be a prevalent event in the development of SCCHN. Mol. Carcinog. 30:26-36, 2001.

Base Sequence↗

PubMed2001

p-Aminophenol-induced liver toxicity: tentative evidence of a role for acetaminophen.

p-Aminophenol (PAP) is a widely used industrial chemical and a metabolite of analgesics, such as acetaminophen (APAP). It was found recently that PAP, a known nephrotoxicant, could cause acute hepatotoxicity in mice but not in rats. The mechanism of hepatotoxicity is not known. The aim of this study was to investigate the role of N-acetylation of PAP to APAP in PAP-induced toxicity. Male C57BL/6 mice injected intraperitoneally (i.p.) with various doses of PAP were sacrificed at 12 hours for measurement of serum glutamic-pyruvic transaminase (GPT) and sorbitol dehydrogenase (SDH) levels and determination of the extent of hepatic nonprotein sulfhydryl (NPSH) and glutathione (GSH) depletion. Plasma levels of APAP and its metabolites were measured by HPLC after PAP administration. p-Aminophenol depleted NPSH in a dose- and time-dependent manner. Depletion of NPSH in mouse liver occurred at PAP doses above 400 mg/kg. Buthionine sulfoximine (BSO), an inhibitor of GSH synthesis, potentiated the PAP-induced hepatotoxicity. Ascorbate, a reducing agent, did not affect PAP-induced hepatotoxicity and NPSH depletion. After PAP treatment, APAP and its sulfate and glucuronide conjugates as well as GSH conjugates (APAP-cysteine and APAP-mercapturate) were detected in the plasma. The results suggest the roles of GSH and N-acetylation of PAP to APAP in PAP-induced hepatotoxicity.

Acetaminophen↗

PubMed2001

Identification of a new gene product of PKIbeta by HPLC-ESIMS.

An identification method using high-performance liquid chromatography combined with electrospray mass spectrometry (HPLC-ESIMS) has been developed to verify an expressed gene product of kinase inhibitor (PKIbeta). This protein was expressed in this university for the first time from a newly cloned gene in the cDNA library of human fetal brain. The measured MW (8468.9 Da) of PKIbeta-78 was consistent with expectations. The gene product of PKIbeta-78 was monitored by ESIMS to ensure there was no mis-expressed PKIbeta-70 in the process of gene engineering. The peptide mapping of PKIbeta-78 and its partial sequence were, furthermore, determined. By database searching based on the experimental MWs and partial sequences provided, it was verified that this gene product is a new protein. The pseudosubstrate site and leucine-rich site for the function region of PKIbeta-78 are also confirmed.

Amino Acid Sequence↗

PubMed2001

A study on the detection of weak photoacoustic signals by stochastic resonance.

On the basis of stochastic resonance (SR) theory a study of the detection of photoacoustic (PA) signals without a lock-in amplifier is presented. The applicability of the SR method is investigated both with simulated signals calculated by use of a Gaussian model and with experimental PA signals. Signal-to-noise ratio (SNR) is improved substantially by adjusting the properties of a non-linear system to realize SR. By use of this method the lock-in amplifier can be removed from a PA spectrometer.

Journal Article↗

PubMed2001

Atrial fibrillation surgery simplified with cryoablation to improve left atrial function.

BACKGROUND: The Maze procedure restores atrial fibrillation to normal sinus rhythm. However, concurrent left atrial functional recovery is not always achieved. To address this limitation, a modification using linear cryoablation is described. METHODS: Between July 1997 and December 1999, 83 patients received atrial fibrillation surgery in association with mitral valve surgery with or without additional concurrent procedures by either the conventional technique, group I (n = 30) or the modified technique, group II (n = 53). Onset of sinus conversion and echocardiographic assessment of postoperative left ventricular function, left atrial size, and mitral A-wave velocity were compared in the early postoperative period and 6 months after surgery. RESULTS: Sinus conversion occurred significantly earlier in group II, 2.4 +/- 5 days versus group I, 7.0 +/- 10 days. The mean transmitral A-wave velocity and the incidence of A-wave appearance in the early postoperative period and 6 months postoperatively were greater in group II than group I. CONCLUSIONS: With the current modification, restoration of sinus rhythm and superior left atrial contractile function occurred earlier than with the standard Maze III technique.

Atrial Fibrillation↗

PubMed2001

Electrical activity modulates growth cone guidance by diffusible factors.

Brief periods of electrical stimulation of cultured Xenopus spinal neurons resulted in a marked alteration in the turning responses of the growth cone induced by gradients of attractive or repulsive guidance cues. Netrin-1-induced attraction was enhanced, and the repulsion induced by myelin-associated glycoprotein (MAG) or myelin membrane fragments was converted to attraction. The effect required the presence of extracellular Ca(2+) during electrical stimulation and appeared to be mediated by an elevation of both cytoplasmic Ca(2+) and cAMP. Thus, electrical activity may influence the axonal path finding of developing neurons, and intermittent electrical stimulation may be effective in promoting nerve regeneration after injury.

Animals↗

PubMed2001

Phosphoprotein-protein interactions revealed by the crystal structure of kinase-associated phosphatase in complex with phosphoCDK2.

The CDK-interacting protein phosphatase KAP dephosphorylates phosphoThr-160 (pThr-160) of the CDK2 activation segment, the site of regulatory phosphorylation that is essential for kinase activity. Here we describe the crystal structure of KAP in association with pThr-160-CDK2, representing an example of a protein phosphatase in complex with its intact protein substrate. The major protein interface between the two molecules is formed by the C-terminal lobe of CDK2 and the C-terminal helix of KAP, regions remote from the kinase-activation segment and the KAP catalytic site. The kinase-activation segment interacts with the catalytic site of KAP almost entirely via the phosphate group of pThr-160. This interaction requires that the activation segment is unfolded and drawn away from the kinase molecule, inducing a conformation of CDK2 similar to the activated state observed in the CDK2/cyclin A complex.

Amino Acid Sequence↗

PubMed2001

The cell biology of neuronal navigation.

Morphogenesis of the nervous system requires the directed migration of postmitotic neurons to designated locations in the nervous system and the guidance of axon growth cones to their synaptic targets. Evidence suggests that both forms of navigation depend on common guidance molecules, surface receptors and signal transduction pathways that link receptor activation to cytoskeletal reorganization. Future challenges remain not only in identifying all the components of the signalling pathways, but also in understanding how these pathways achieve signal amplification and adaptation-two essential cellular processes for neuronal navigation.

Animals↗

PubMed2001

Sequence-specific termination by T7 RNA polymerase requires formation of paused conformation prior to the point of RNA release.

BACKGROUND: The sequence-specific, hairpin-independent termination signal for the bacteriophage RNA polymerases in Escherichia coli rrnB t1 terminator consists of two modules. The upstream module includes the conserved sequence and the downstream one is U-rich. RESULTS: Elongation complexes of T7 RNA polymerase paused 2 bp before reaching the termination site at a 500 microM concentration of NTP. At 5-50 microM NTP, however, they paused and terminated there or resumed elongation beyond the termination site. Only at higher concentrations of NTP (500 microM), the pause complex proceeded slowly to and became incompetent at the termination site. At 4 bp or more before the termination site, the unprotected single-stranded region of transcription bubble shrank at the trailing edge to 4-5 bp from approximately 10 bp, resulting from duplex formation of the conserved sequence. The pause and bubble collapse were not observed with an inactive mutant of the termination signal. CONCLUSION: Sequence-specific termination requires the slow elongation mode of paused conformation, working only at high concentrations of NTP for a few bp prior to the RNA release site. The collapse of bubble that was observed several base pairs before the termination site and/or the resulting duplex might subsequently lead to the paused conformation of T7 elongation complexes.

Bacteriophage T7↗

PubMed2001

Comparison of the IAEA TRS-398 and AAPM TG-51 absorbed dose to water protocols in the dosimetry of high-energy photon and electron beams.

The International Atomic Energy Agency (IAEA TRS-398) and the American Association of Physicists in Medicine (AAPM TG-51) have published new protocols for the calibration of radiotherapy beams. These protocols are based on the use of an ionization chamber calibrated in terms of absorbed dose to water in a standards laboratory's reference quality beam. This paper compares the recommendations of the two protocols in two ways: (i) by analysing in detail the differences in the basic data included in the two protocols for photon and electron beam dosimetry and (ii) by performing measurements in clinical photon and electron beams and determining the absorbed dose to water following the recommendations of the two protocols. Measurements were made with two Farmer-type ionization chambers and three plane-parallel ionization chamber types in 6, 18 and 25 MV photon beams and 6, 8, 10, 12, 15 and 18 MeV electron beams. The Farmer-type chambers used were NE 2571 and PTW 30001, and the plane-parallel chambers were a Scanditronix-Wellhöfer NACP and Roos, and a PTW Markus chamber. For photon beams, the measured ratios TG-51/TRS-398 of absorbed dose to water Dw ranged between 0.997 and 1.001, with a mean value of 0.999. The ratios for the beam quality correction factors kQ were found to agree to within about +/-0.2% despite significant differences in the method of beam quality specification for photon beams and in the basic data entering into kQ. For electron beams, dose measurements were made using direct N(D,w) calibrations of cylindrical and plane-parallel chambers in a 60Co gamma-ray beam, as well as cross-calibrations of plane-parallel chambers in a high-energy electron beam. For the direct N(D,w) calibrations the ratios TG-51/TRS-398 of absorbed dose to water Dw were found to lie between 0.994 and 1.018 depending upon the chamber and electron beam energy used, with mean values of 0.996, 1.006, and 1.017, respectively, for the cylindrical, well-guarded and not well-guarded plane-parallel chambers. The Dw ratios measured for the cross-calibration procedures varied between 0.993 and 0.997. The largest discrepancies for electron beams between the two protocols arise from the use of different data for the perturbation correction factors p(wall) and p(dis) of cylindrical and plane-parallel chambers, all in 60Co. A detailed analysis of the reasons for the discrepancies is made which includes comparing the formalisms, correction factors and the quantities in the two protocols.

Air↗

PubMed2001

Genetic control of horizontal virus transmission in the chestnut blight fungus, Cryphonectria parasitica.

Vegetative incompatibility in fungi has long been known to reduce the transmission of viruses between individuals, but the barrier to transmission is incomplete. In replicated laboratory assays, we showed conclusively that the transmission of viruses between individuals of the chestnut blight fungus Cryphonectria parasitica is controlled primarily by vegetative incompatibility (vic) genes. By replicating vic genotypes in independent fungal isolates, we quantified the effect of heteroallelism at each of six vic loci on virus transmission. Transmission occurs with 100% frequency when donor and recipient isolates have the same vic genotypes, but heteroallelism at one or more vic loci generally reduces virus transmission. Transmission was variable among single heteroallelic loci. At the extremes, heteroallelism at vic4 had no effect on virus transmission, but transmission occurred in only 21% of pairings that were heteroallelic at vic2. Intermediate frequencies of transmission were observed when vic3 and vic6 were heteroallelic (76 and 32%, respectively). When vic1, vic2, and vic7 were heteroallelic, the frequency of transmission depended on which alleles were present in the donor and the recipient. The effect of heteroallelism at two vic loci was mostly additive, although small but statistically significant interactions (epistasis) were observed in four pairs of vic loci. A logistic regression model was developed to predict the probability of virus transmission between vic genotypes. Heteroallelism at vic loci, asymmetry, and epistasis were the dominant factors controlling transmission, but host genetic background also was statistically significant, indicating that vic genes alone cannot explain all the variation in virus transmission. Predictions from the logistic regression model were highly correlated to independent transmission tests with field isolates. Our model can be used to estimate horizontal transmission rates as a function of host genetics in natural populations of C. parasitica.

Alleles↗

PubMed2001

Connection between the inner ear and the lymphatic system.

OBJECTIVE: The aim of this study was to identify the lymphatic drainage of the inner ear in guinea pigs. STUDY DESIGN: Prospective study. METHODS: The prospective study was performed in guinea pigs by injection of keyhole limpet hemocyanin (KLH) into either the right-side scala tympani or the middle ear cavity. The left side was not injected and served as a control. Fifteen minutes after injection, the animals were killed by intracardiac perfusion with paraformaldehyde and tissue specimens (right and left temporal bones, cervical lymph nodes, and the spleen) were collected. The presence of KLH in each specimen was determined by immunohistochemical assay of frozen sections using polyclonal mouse anti-KLH antibodies. RESULTS: After injection into the middle ear, labeled cells were identified in the parotid, superficial ventral, mandibular, and deep cranial cervical lymph nodes. However, after inner ear injections KLH was present in only the parotid and superficial ventral cervical nodes. The spleen contained KLH-positive cells following injection into either the middle or inner ear, but not all animals contained labeled spleen cells. CONCLUSIONS: The inner ear has a connection to the lymphatic drainage system. Because fewer lymph nodes contained labeled cells after inner ear injection than after middle ear injection, it is concluded that the inner ear does not simply drain to the middle ear and subsequently to the lymph nodes but seems likely to have its own connections.

Animals↗