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Biomedical subjects

H Tu

Publications and source records attributed to H Tu.

At least 37 records · Page 2Linked to original sources
PubMed2000

A novel electrochemical microsensor for the determination of NO and its application to the study of the NO donor S-nitrosoglutathione.

A novel electrochemical microsensor for the determination of NO based on an electropolymerized film of tetraaminophthalocyaninecopper [Cu(TAPc)] was prepared. Its response to NO and its application to the study of an NO donor (S-nitrosoglutathione; GSNO) are also described. The microsensor exhibited an electrocatalytic effect on NO oxidation and showed a low detection limit, high sensitivity and selectivity for NO determination. The oxidation current (measured by differential pulse amperometry) was linear for NO concentrations ranging from 6.2 x 10(-9) to 3.0 x 10(-5) mol L-1 with a calculated detection limit of 4.0 x 10(-9) mol L-1 (S/N = 3) and a linear coefficient of 0.9984. Some endogenous electroactive substances in biological tissues, such as dopamine, 5-hydroxytryptamine and nitrite, at concentrations higher than those in biological systems did not interfere with NO determination. The sensor shows promise for the possible in vivo determination of NO. Using the microsensor, the NO release from the NO donor (GSNO) was successfully monitored. This work sets a foundation for the study of the pharmacology and the biological effects in vivo of S-nitrosothiols.

Electrochemistry↗
PubMed2000

Does type of health insurance affect health care use and assessments of care among the privately insured?

OBJECTIVE: To inform the debate about managed care by examining how different types of private insurance-indemnity insurance, PPOs, open model HMOs, and closed model HMOs-affect the use of health services and consumer assessments of care. DATA SOURCES/DATA COLLECTION: The 1996-1997 Community Tracking Study Household Survey, a nationally representative telephone survey of households, and the Community Tracking Study Insurance Followback Survey, a supplement to the Household Survey, which asks insurance organizations to match household respondents to specific insurance products. The analysis sample includes 27,257 nonelderly individuals covered by private insurance. STUDY DESIGN: Based on insurer reports, individuals are grouped into one of the four insurance product types. Measures of service use include ambulatory visits, preventive care use, hospital use, surgeries, specialist use, and whether there is a usual source of care. Consumer assessments of care include unmet or delayed care needs, satisfaction with health care, ratings of the last physician visit, and trust in physicians. Estimates are adjusted to control for differences in individual characteristics and location. PRINCIPAL FINDINGS: As one moves from indemnity insurance to PPOs to open model HMOs to closed model HMOs, use of primary care increases modestly but use of specialists is reduced. Few differences are observed in other areas of service use, such as preventive care, hospital use, and surgeries. The likelihood of having unmet or delayed care does not vary by insurance type, but the reasons that underlie such access problems do vary: enrollees in more managed products are less likely to cite financial barriers to care but are more likely to perceive problems in provider access, convenience, and organizational factors. Consumer assessments of care-including satisfaction with care, ratings of the last physician visit, and trust in physicians-are generally lower under more managed products, particularly closed model HMOs. CONCLUSIONS: The type of insurance that people have-not just whether it is managed care but the type of managed care-affects their use of services and their assessments of the care they receive. Consumers and policymakers should be reminded that managed care encompasses a variety of types of insurance products that have different effects and may require different policy responses.

Adult↗
PubMed1999

Hepatitis C virus RNA polymerase and NS5A complex with a SNARE-like protein.

Hepatitis C virus (HCV) NS5A is a phosphoprotein that possesses a cryptic trans-activation activity. To investigate its potential role in viral replication, we searched for the cellular proteins interacting with NS5A protein by yeast two-hybrid screening of a human hepatocyte cDNA library. We identified a newly discovered soluble N-ethylmaleimide-sensitive factor attachment protein receptor-like protein termed human vesicle-associated membrane protein-associated protein of 33 kDa (hVAP-33). In vitro binding assay and in vivo coimmunoprecipitation studies confirmed the interaction between hVAP-33 and NS5A. Interestingly, hVAP-33 was also shown to interact with NS5B, the viral RNA-dependent RNA polymerase. NS5A and NS5B bind to different domains of hVAP-33: NS5A binds to the C-terminus, whereas NS5B binds to the N-terminus of hVAP-33. Immunofluorescent staining showed a significant colocalization of hVAP-33 with both NS5A and NS5B proteins. hVAP-33 contains a coiled-coil domain followed by a membrane-spanning domain at its C-terminus. Cell fractionation analysis revealed that hVAP-33 is predominantly associated with the ER, the Golgi complex, and the prelysosomal membrane, consistent with its potential role in intracellular membrane trafficking. These interactions provide a mechanism for membrane association of the HCV RNA replication complex and further suggest that NS5A is a part of the viral RNA replication complex.

Animals↗
PubMed1999

Evidence for 4-hydroxyproline in viral proteins. Characterization of a viral prolyl 4-hydroxylase and its peptide substrates.

4-Hydroxyproline, the characteristic amino acid of collagens and collagen-like proteins in animals, is also found in certain proline-rich proteins in plants but has been believed to be absent from viral and bacterial proteins. We report here on the cloning and characterization from a eukaryotic algal virus, Paramecium bursaria Chlorella virus-1, of a 242-residue polypeptide, which shows distinct sequence similarity to the C-terminal half of the catalytic alpha subunits of animal prolyl 4-hydroxylases. The recombinant polypeptide, expressed in Escherichia coli, was found to be a soluble monomer and to hydroxylate both (Pro-Pro-Gly)(10) and poly(L-proline), the standard substrates of animal and plant prolyl 4-hydroxylases, respectively. Synthetic peptides such as (Pro-Ala-Pro-Lys)(n), (Ser-Pro-Lys-Pro-Pro)(5), and (Pro-Glu-Pro-Pro-Ala)(5) corresponding to proline-rich repeats coded by the viral genome also served as substrates. (Pro-Ala-Pro-Lys)(10) was a particularly good substrate, with a K(m) of 20 microM. The prolines in both positions in this repeat were hydroxylated, those preceding the alanines being hydroxylated more efficiently. The data strongly suggest that P. bursaria Chlorella virus-1 expresses proteins in which many prolines become hydroxylated to 4-hydroxyproline by a novel viral prolyl 4-hydroxylase.

Amino Acid Sequence↗
PubMed1999

Identification of a nuclear receptor for bile acids.

Bile acids are essential for the solubilization and transport of dietary lipids and are the major products of cholesterol catabolism. Results presented here show that bile acids are physiological ligands for the farnesoid X receptor (FXR), an orphan nuclear receptor. When bound to bile acids, FXR repressed transcription of the gene encoding cholesterol 7alpha-hydroxylase, which is the rate-limiting enzyme in bile acid synthesis, and activated the gene encoding intestinal bile acid-binding protein, which is a candidate bile acid transporter. These results demonstrate a mechanism by which bile acids transcriptionally regulate their biosynthesis and enterohepatic transport.

Animals↗
PubMed1999

Genetic evidence for Pak1 autoinhibition and its release by Cdc42.

Pak1 protein kinase of Schizosaccharomyces pombe, a member of the p21-GTPase-activated protein kinase (PAK) family, participates in signaling pathways including sexual differentiation and morphogenesis. The regulatory domain of PAK proteins is thought to inhibit the kinase catalytic domain, as truncation of this region renders kinases more active. Here we report the detection in the two-hybrid system of the interaction between Pak1 regulatory domain and the kinase catalytic domain. Pak1 catalytic domain binds to the same highly conserved region on the regulatory domain that binds Cdc42, a GTPase protein capable of activating Pak1. Two-hybrid, mutant, and genetic analyses indicated that this intramolecular interaction rendered the kinase in a closed and inactive configuration. We show that Cdc42 can induce an open configuration of Pak1. We propose that Cdc42 interaction disrupts the intramolecular interactions of Pak1, thereby releasing the kinase from autoinhibition.

Amino Acid Sequence↗
PubMed1999

Dauricine suppressed CsCl-induced early afterdepolarizations and triggered arrhythmias in rabbit heart in vivo.

AIM: To study the effect of dauricine on CsCl-induced early afterdepolarizations (EAD) and ventricular arrhythmias in rabbits. METHODS: Monophasic action potentials (MAP) of the left ventricle of the rabbit heart in situ were recorded with MAP recording technique. CsCl 1-2 mmol.kg-1 i.v. was used to induce EAD and ventricular arrhythmias. RESULTS: CsCl resulted in decrease of MAP amplitude (MAPA, P < 0.05) and prolongation of MAP duration at 90% repolarization (MAPD90, P < 0.01), QRS, and R-R duration (P < 0.05) compared with those before CsCl in the dauricine and control group. CsCl injection induced EAD that appeared within about 30 s and disappeared 5-15 min thereafter. EAD always preceded ventricular arrhythmias including ventricular premature beats and paroxysmal ventricular tachycardia. The EAD amplitude (EADA) in the dauricine group (26% +/- 9% of MAPA) was smaller than that in the control group (52% +/- 5% of MAPA, P < 0.05) and the incidence of arrhythmias in dauricine group (28%) was lower than that in control group (80%, P < 0.05). CONCLUSION: Dauricine exerted an antagonistic effect on EAD and suppressed triggered ventricular arrhythmias by decreasing EADA.

Action Potentials↗
PubMed1997

Frequency of hepatitis B virus e-minus mutants varies among patients from different areas of China.

Four hundred forty-six serum samples from HBsAg-positive chronic hepatitis B patients were collected from five areas in China (eastern coastal city, Shanghai; southwestern inland city, Chengdu; mid-inland city, Wuhan; southern island city, Haikou; and northeastern city, Changchun). Precore stop codon variants (e-minus mutants) were screened using a rapid method of polymerase chain reaction (PCR) amplification of a precore and partial core gene fragment (nucleotides 1785-2172), followed by dot-blot hybridization with specific oligonucleotide probes (M0, and M1 + M2). The sequence of the M0 probe covered the distal precore region of wild-type virus (nucleotides 1887-1908), and the sequences of the M1 and M2 probes were from sequences mutated at nt. 1898, (TGG-->TAG) with or without additional change at nt. 1901. A significantly lower incidence of the precore stop codon was found in anti-HBe-positive serum samples from Haikou (17.6%), whereas in other areas the percentages of this mutation in anti-HBe positive sera ranged from 47.4% to 78.9%. In HBeAg-positive samples, the rate of e-minus mutant in coexistence with wild-type virus was low in specimens from Haikou (9.5%) and Changchun (2.9%) compared to other areas in China. In contrast, coexistence of mutant and wild-type virus was frequently detected in samples from Wuhan (50.0%).

China↗
PubMed1997

Multiple regulatory domains on the Byr2 protein kinase.

Byr2 protein kinase, a homolog of mammalian mitogen-activated protein kinase/extracellular signal-regulated kinase kinase (MEKK) and Saccharomyces cerevisiae STE11, is required for pheromone-induced sexual differentiation in the fission yeast Schizosaccharomyces pombe. Byr2 functions downstream of Ste4, Ras1, and the membrane-associated receptor-coupled heterotrimeric G-protein alpha subunit, Gpa1. Byr2 has a distinctive N-terminal kinase regulatory domain and a characteristic C-terminal kinase catalytic domain. Ste4 and Ras1 interact with the regulatory domain of Byr2 directly. Here, we define the domains of Byr2 that bind Ste4 and Ras1 and show that the Byr2 regulatory domain binds to the catalytic domain in the two-hybrid system. Using Byr2 mutants, we demonstrate that these direct physical interactions are all required for proper signaling. In particular, the physical association between Byr2 regulatory and catalytic domains appears to result in autoinhibition, the loss of which results in kinase activation. Furthermore, we provide evidence that Shk1, the S. pombe homolog of the STE20 protein kinase, can directly antagonize the Byr2 intramolecular interaction, possibly by phosphorylating Byr2.

Cloning, Molecular↗
PubMed1997

[Mutations and their significance in the corepromoter region of hepatitis B virus].

OBJECTIVE: To study the mutations in the core promoter (CP) region of hepatitis B virus (HBV) in Chinese viral hepatits B patients. METHODS: CP regions of 48 HBV strains were analysed by polymerase chain reaction (PCR) direct sequencing approach. RESULTS: 67% of the samples were detected to have point mutations in the CP region. The hot spots located in nt 1754-1766 and nt 1801-1811 while nucleotides in nt 1777-1800 and nt 1812-1836 were highly conserved. nt. 1764 mutation was present in HBeAg negative patients. There were 16 point mutations in region overlapping X gene, 9 of which lead to amino acid change. CONCLUSION: Though mutations in CP region of HBV appear frequently, the sequences associated with viral transcription are rarely changed. Point mutation at nt 1764 is related to HBeAg negative phenotype, but it is not the specific mutation of fulminant hepatitis. The importance of mutations in the X gene overlapping region needs to be further investigated.

Genetic Variation↗
PubMed1996

Incidence of apoptosis and cell proliferation in prostate cancer: relationship with TGF-beta1 and bcl-2 expression.

The incidence of programmed cell death (apoptosis) and cell proliferation was investigated in the normal and malignant human prostate to define the significance of their potential deregulation in human prostate cancer. The incidence of "spontaneous" apoptosis was analyzed using an in situ end-labeling procedure for detection of nucleosomal DNA fragmentation, as well as the pattern and topological localization of expression of the 2 proteins regulating apoptosis, TGF-beta1, and bcl-2, in 40 primary prostatic adenocarcinomas with varying tumor grades, 17 lymph nodes positive for metastatic prostate cancer and 9 normal prostate specimens. The basal level of cell proliferation of the different prostatic cell populations in the same specimens was determined, utilizing the Ki-67 nuclear antigen. Localized prostate cancer cells exhibited a relatively low rate of apoptosis, which was significantly lower than the apoptotic index of normal prostate glandular epithelial cells. Metastatic prostate tumor cells, however, exhibited a significantly higher apoptotic index compared with localized prostate cancer cells. A significant increase in the proliferative index was detected in prostatic tumors compared with the normal gland (5-fold), and there was an even more marked elevation in the proliferative index of the metastatic prostate tumor cells compared to the normal prostate epithelial cells (approximately 24-fold). Immunohistochemical analysis of normal and malignant prostate specimens revealed a predominant TGF-beta immunoreactivity in the glandular epithelial cells, while the stromal component was totally negative. There was a significant increase in the levels of TGF-beta in primary prostatic tumors compared to the normal prostate. Bcl-2 expression was detected among certain populations of tumor epithelial cells in a mutually exclusive topological distribution pattern for apoptosis. In marked contrast, neither TGF-beta1 nor bcl-2 immunoreactivity was detected in metastatic prostate tumor cells, despite their high proliferative and apoptotic rates. Balancing the prostatic growth equation for the prostatic tumor epithelial cell populations revealed a substantial net increase in cell number in both primary and metastatic prostate cancers. This loss of apoptotic control in favor of cell proliferation may be responsible for prostate cancer initiation and progression.

Adenocarcinoma↗
PubMed1996

Type 2 iodothyronine deiodinase is highly expressed in human thyroid.

Type 2 iodothyronine deiodinase (D2) is a recently cloned selenodeiodinase thought to provide intracellular 3,5,3' triiodothyronine (T3) to a restricted group of tissues. We report here the presence of D2 mRNA in human thyroid at levels 50-150-fold higher than in placenta. Surprisingly, while type 1 deiodinase (D1) is known to be present in human thyroid, D2 has not been evaluated previously. D2 mRNA was especially high in thyroids from Graves' patients and in follicular adenomas. Stimulated thyroids had higher D2 to D1 mRNA ratios than normal or multinodular glands suggesting differential regulation of D1 and D2 expression. Microsomes from normal, Graves', and TSH-stimulated thyroids contained low Km D2 activity resistant to propylthiouracil (1 mM) or to inactivation by N-bromoacetyl T3, agents which block or inactivate D1. At 2 nM thyroxine (T4), 100 times the physiological-free T4 levels, 60-80% of T4 to T3 conversion in stimulated, but only 27% of that in normal thyroids, is catalyzed by D2. We conclude that intrathyroidal T4 to T3 conversion by D2 may contribute significantly to the relative increase in thyroidal T3 production in patients with Graves' disease, toxic adenomas, and, perhaps, iodine deficiency.

Enzyme Inhibitors↗
PubMed1996

Apoptotic versus proliferative activities in human benign prostatic hyperplasia.

Cell growth in the normal prostate is regulated by a delicate balance between cell death and cell proliferation (ie, apoptotic v proliferative activity). Disruption of the molecular mechanisms that regulate these two processes may underline the abnormal growth of the gland leading to benign prostatic hyperplasia (BPH). In this study, the incidence of programmed cell death (apoptosis) and cell proliferation was comparatively analyzed among the various cell subpopulations in the normal and benign hyperplastic human prostate. The authors also examined the relative expression of two proteins involved in the regulation of prostate apoptosis: (1) transforming growth factor (TGF)-beta1, a negative growth factor able to induce prostate apoptosis under physiological conditions; and (2) bcl-2, a potent apoptosis suppressor. Analysis of the incidence of "spontaneous" apoptosis in situ, using the end-labeling terminal transferase staining technique for the detection of nucleosomal DNA fragmentation, revealed infrequent apoptotic staining in isolated basal and secretory prostate epithelial cells. The basal level of cell proliferation was determined on the basis of the Ki-67 nuclear antigen staining, a nuclear protein that appears primarily during the proliferative phases of the cell cycle. The Ki-67-positive nuclei were equally distributed among the basal and secretory epithelial cells of the hyperplastic prostatic acini. The apoptotic index of the secretory and basal cells of the prostate epithelium was higher in the normal prostate compared with BPH tissue, whereas there was a significant increase in the proliferative index of the respective cell populations in the hyperplastic prostate. Balancing the apoptotic versus the proliferative activities revealed a substantial net decrease (fourfold) in the total number of cells dying via apoptosis in both the glandular and basal epithelial cell compartments of the hypertrophic prostate (BPH) when compared with the normal gland. TGF-beta staining was exclusively identified in the secretory epithelial cells, lining the prostatic lumen with minimal involvement of the basal cells and total lack of immunoreactivity among the stroma elements. Statistical analysis revealed a significant elevation in TGF-beta expression in the epithelial cells of BPH tissue compared with the normal prostate (P < .001). Expression of bcl-2 was topologically restricted to the glandular epithelium of the prostate. In the normal prostate, bcl-2 immunoreactivity was predominantly identified in the basal cell layer. An increase in both the intensity of immunoreactivity for bcl-2 and the number of positive epithelial cells (basal and secretory) was detected in BPH specimens relative to the normal prostate (P < .02). These results suggest a potential involvement of enhanced expression of this antiapoptosis protein in deregulation of the normal apoptotic cell death mechanisms in the human prostate, thus resulting in a growth imbalance in favor of cell proliferation that might ultimately promote prostatic hyperplasia.

Adult↗
PubMed1996

Identification of Ste4 as a potential regulator of Byr2 in the sexual response pathway of Schizosaccharomyces pombe.

A conserved MAP kinase cascade is central to signal transduction in both simple and complex eukaryotes. In the yeast Schizosaccharomyces pombe, Byr2, a homolog of mammalian MAPK/ERK kinase kinase and Saccharomyces cerevisiae STE11, is required for pheromone-induced sexual differentiation. A screen for S. pombe proteins that interact with Byr2 in a two-hybrid system led to the isolation of Ste4, a protein that is known to be required for sexual function. Ste4 binds to the regulatory region of Byr2. This binding site is separable from the binding site for Ras1. Both Ste4 and Ras1 act upstream of Byr2 and act at least partially independently. Ste4 contains a leucine zipper and is capable of homotypic interaction. Ste4 has regions of homology with STE50, an S. cerevisiae protein required for sexual differentiation that we show can bind to STE11.

Binding Sites↗
PubMed1995

Two microtubule-associated proteins required for anaphase spindle movement in Saccharomyces cerevisiae.

In many eucaryotic cells, the midzone of the mitotic spindle forms a distinct structure containing a specific set of proteins. We have isolated ASE1, a gene encoding a component of the Saccharomyces cerevisiae spindle midzone. Strains lacking both ASE1 and BIK1, which encodes an S. cerevisiae microtubule-associated protein, are inviable. The analysis of the phenotype of a bik1 ase1 conditional double mutant suggests that BIK1 and ASE1 are not required for the assembly of a bipolar spindle, but are essential for anaphase spindle elongation. The steady-state levels of Ase1p are regulated in a manner that is consistent with a function during anaphase: they are low in G1, accumulate to maximal levels after S phase and then drop as cells exit mitosis. Components of the spindle midzone may therefore be required in vivo for anaphase spindle movement. Additionally, anaphase spindle movement may depend on a dedicated set of genes whose expression is induced at G2/M.

Anaphase↗
PubMed1994

Rare incidence of human papillomavirus types 16 and 18 in primary and metastatic human prostate cancer.

OBJECTIVES: The reported data regarding the incidence and significance of human papillomavirus (HPV) in prostate cancer have been inconsistent. In the present study the incidence of HPV 16 and 18 was evaluated in an expanded series of primary as well as metastatic prostate cancer specimens, in order to evaluate a potential role of HPV infection in development and progression of prostate cancer. This is the first study attempting to establish the significance of HPV in metastatic prostate cancer. METHODS: The presence of high risk human papillomaviruses HPV 16 and 18 was analyzed using the polymerase chain reaction (PCR) amplification method and Southern blot hybridization analysis in a total of 61 prostatic tissue specimens: 43 primary prostate adenocarcinoma formalin-fixed, paraffin-embedded specimens, with varying degrees of differentiation (mean Gleason score 5.8, range 3 to 9); 17 pelvic lymph nodes positive for metastatic deposits; and 1 normal prostate specimen. RESULTS: This human papillomavirus typing indicates that only 1 out of the 43 prostatic specimens analyzed was positive for HPV 16 and 1 metastatic lymph node was positive for HPV 18, as revealed by Southern analysis. These results demonstrate the infrequent detection of HPV 16 and 18 DNA in all the primary prostatic adenocarcinoma specimens and metastatic lymph nodes analyzed in this study population. CONCLUSIONS: The negative HPV status for primary and metastatic prostate cancer demonstrated in this study provides a strong argument against an etiological role of HPV infection in the development and progression of the disease.

Adenocarcinoma↗