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PubMed1999

Ethanol consumption by Fawn-Hooded rats following abstinence: effect of naltrexone and changes in mu-opioid receptor density.

BACKGROUND: Relapse after abstinence can be modelled in rats using an alcohol deprivation effect (ADE) of enhanced ethanol consumption after a period of enforced abstinence from ethanol; however, not all rat strains display such an effect. We wanted to examine the effect of naltrexone on ethanol consumption by ethanol-preferring Fawn-Hooded (FH) rats using such a model. METHODS: FH rats were given continual free-choice access to a 5% ethanol solution or water (4 weeks) followed by 2 weeks of water alone. At the end of this abstinence period, osmotic minipumps were implanted subcutaneously to deliver saline (n = 4) or naltrexone (n = 4; 8.4 mg/kg/day for 4 weeks). After recovery from surgery, the rats were again given access to 5% ethanol under the same free-choice conditions (4 weeks). A third group of age-matched controls drank only water during the behavioral trial. At the end of the behavioral trial, the rats were decapitated and an autoradiographic examination was made of micro-opioid receptor density through the forebrain using the ligand [125I]FK-33824. RESULTS: First, a period of enforced abstinence from ethanol consumption caused a significant (p < 0.05) and prolonged increase in ethanol preference (+18%) and decrease in water consumption (-53%), although the volume of ethanol consumed (ml/day) did not vary, indicating an atypical ADE in this rat strain. Second, naltrexone significantly (p < 0.05) decreased ethanol consumption by the FH rats in terms of absolute amount of ethanol consumed and preference for ethanol solution, but this effect of naltrexone diminished over time, concurrent with a robust and significant elevation in micro-opioid receptor density in all brain regions examined (p < 0.05). Finally, ethanol consumption alone also upregulated micro-opioid receptor density relative to nondrinking controls in a number of brain regions, which included the nucleus accumbens (+29%) and caudate-putamen (+15%,p < 0.05), but decreased micro-opioid receptor density in other regions including the substantia nigra pars reticulata, which was suggestive of an indirect effect on micro-opioid receptors. CONCLUSIONS: The data suggest that continual long-term naltrexone treatment may not be effective in the treatment of alcoholism, possibly because of the induced increase in micro-opioid receptor density.

Alcohol Drinking↗

PubMed1999

Use of a mail-out continuing education article to teach health professionals about drug-induced disease.

A U.S. Food and Drug Administration (FDA)/Georgetown University Medical Center conference was the basis for "Clinical Therapeutics and the Recognition of Drug-Induced Disease," the first MEDWATCH continuing education (CE) mail-out article. Developed as a major component of FDA MEDWATCH post-marketing surveillance outreach, the article used a clinical therapeutic approach to discuss topics including adverse drug events (ADEs) pharmacology and ADE reporting. Distributed nationwide through the MEDWATCH Partners, health professionals applied for CE credit by completing a self-assessment examination. With the overall response rate slightly more than 2%, 15,260 health professionals (55% physicians and 37% pharmacists) received CE credit. Evaluation of the initial approximately two-thirds (N = 10,021) of successfully completed exams found 99% agreement that stated learning objectives were met, and the article relevant to their clinical practice; spontaneous comments/letters were also very positive. The highest percentage responding specialists were internists (28%) and psychiatrists (17%), with notable differences found among specialties for response rate versus relative article distribution (such as relatively low response rates among surgeons and radiology/radiation physics specialists). The number of health professionals receiving CE credit, coupled with examination performance and overall response, indicates that "Clinical Therapeutics and the Recognition of Drug-Induced Disease" was well received and fulfilled learning objectives. The results provide encouragement for this continuing educational approach.

Adverse Drug Reaction Reporting Systems↗

PubMed2001

Age-related radical-induced DNA damage is linked to prostate cancer.

We measured concentrations and ratios of mutagenic (8-OH) lesions to putatively nonmutagenic formamidopyrimidine (Fapy) lesions of adenine (Ade) and guanine (Gua) to elucidate radical (.OH)-induced changes in DNA of normal, normal from cancer, and cancer tissues of the prostate. The relationship between the lesions was expressed using the mathematical model log(10)[(8-OH-Ade + 8-OH-Gua)/(FapyAde + FapyGua)]. Logistic regression analysis of the log ratios for DNA of normal and cancer tissues discriminated between the two tissue groups with high sensitivity and specificity. Correlation analysis of log ratios for normal prostates revealed a highly significant increase in the proportion of mutagenic base lesions with age. Data from correlation analysis of the log ratios for normal tissues from cancer were consistent with an age-dependent, dose-response relationship. The slopes for both correlations intersected at approximately 61 years, an age when prostate cancer incidence is known to rise sharply. The age-related increase in the proportion of.OH-induced mutagenic base lesions is likely a significant factor in prostate cancer development.

Adenine↗

PubMed1975

Hypervitaminosis A in the dog.

Toxic effects of excessive vitamin A and of excessive vitamin ADE were studied in 9 mixed Labrador Retriever pups. Clinical signs were loss of body weight, dullness, emaciation, roughened coat, evident pain in limb joints, and retarded growth. Radiologic changes were decreases in overall length and thickness of long bones, development of osteophytes, periosteal reaction, and premature closure of epiphyses. Pathologic changes were degenerative epiphyseal plate, hemorrhage and exostotic proliferation of periosteum, fatty liver, and microcalculi in kidney. Toxic effects of excessive vitamin A did not appear to be so great when it was administered as vitamin ADE.

Animals↗

PubMed2003

[Screening of the genes of hepatitis B virus PreS2 interacting proteins].

OBJECTIVE: To screen and clone the genes of proteins in hepatocytes interacting with hepatitis B virus (HBV) PreS2 by yeast-two hybridization technique. METHODS: The HBV PreS2 gene was amplified by polymerase chain reaction (PCR) and HBV PreS2 bait plasmid was constructed by using yeast-two hybridization system 3, then transformed into yeast AH109, followed by mating with yeast Y187 containing liver cDNA library plasmid in 2 YPDA medium. Diploid yeast was plated on synthetic dropout nutrient medium (SD/-Trp-Leu-Ade-His) and synthetic dropout nutrient medium (SD/-Trp-Leu-Ade-His) containing X-alpha-gal for selecting positive blue clones, then amplified by PCR, sequenced, and performed bioinformatics analysis. RESULTS: HBV PreS2 gene was cloned successfully and expressed in yeast AH109.Twenty-six positive colonies were selected, among them, twelve containing metallothionein 2A, one cytochrome C oxidase II, two cytochrome P450 subfamily IV4F, two cytochrome c oxidase subunit 4 isoform 1, three albumin (ALB), one Na(+)K(+) transporting ATPase beta-1 polypeptide, two prealbumin, one lectin galactoside-binding subunit, and Two new genes with unknown function. CONCLUSION: Genes of HBV PreS2 interacting proteins have been successfully cloned, which brings some new clues for studying the biological functions of HBV PreS2 and related proteins.

Cloning, Molecular↗

PubMed2003

Changes in water quality parameters due to in-sewer processes.

Combined sewer systems contain a large number of organic and inorganic pollutants from both domestic and industrial sources. These pollutants are often retained within the combined sewer system for significant lengths of time before entering sewage treatment works, or being spilt to a watercourse via a combined sewer overflow (CSO) during storm conditions. Currently little knowledge exists concerning the effects of in sewer processes on pollutants. Understanding of in-sewer processes is important for the effective and efficient design of treatment works and CSO chambers and for impact assessments on receiving waters. A series of studies covering storm and dry weather flow conditions were undertaken with the aim of investigating the nature of in-sewer processes. These studies consisted of marking a body of water with a fluorescent tracer. The tracer was then monitored at a series of downstream sites, and discrete samples collected from the body of water as it progressed through the sewer. The samples were analysed for water quality parameters and these results investigated in tandem with the detailed hydraulic information gained through the tracer studies. The results highlight the hydraulic differences between storm and dry weather conditions such as increased travel times and mixing under storm conditions. The Advection Dispersion Equation (ADE) and Aggregated Dead Zone (ADZ) model parameters have been quantified for the tracer data. The ADE mixing coefficient is shown to increase by an order of magnitude for storm conditions. The ADZ dispersive fraction parameter is shown to be approximately constant with flow. Chemical reactions and decay within the sewer system were found to be consistent with oxygen limitation.

Environmental Monitoring↗

PubMed1992

Regulation of purine nucleotide synthesis in human B lymphoblasts with both hypoxanthine-guanine phosphoribosyltransferase deficiency and phosphoribosylpyrophosphate synthetase superactivity.

Human B lymphoblast lines severely deficient in hypoxanthine-guanine phosphoribosyltransferase (HGPRT) were selected for resistance to 6-thioguanine from cloned normal and phosphoribosylpyrophosphate (PP-Rib-P) synthetase-superactive cell lines and were compared with their respective parental cell lines with regard to growth and PP-Rib-P and purine nucleotide metabolism. During blockade of purine synthesis de novo with 6-methylthioinosine or aminopterin, inhibition of growth of all HGPRT-deficient cell lines was refractory to addition of Ade at concentrations which restored substantial growth to parental cell lines. Ade-resistant inhibition of growth of parental lines by 6-methylthioinosine, however, occurred during Ado deaminase inhibition. Insufficient generation of IMP (and ultimately guanylates) to support growth of lymphoblasts lacking HGPRT activity and blocked in purine synthesis de novo best explained these findings, implying that a major route of interconversion of AMP to IMP involves the reaction sequence: AMP----Ado----Ino----Hyp----IMP. PP-Rib-P generation and purine nucleoside triphosphate pools were unchanged by introduction of HGPRT deficiency into normal lymphoblast lines, in agreement with the view that accelerated purine synthesis de novo in this deficiency results from increased availability of PP-Rib-P for the pathway. Cell lines with dual enzyme defects did not differ from PP-Rib-P synthetase-superactive parental lines in rates of PP-Rib-P and purine synthesis despite 5-6-fold increases in PP-Rib-P concentrations, excretion of nearly 50% of newly synthesized purines, and diminished GTP concentrations. Fixed rates of purine synthesis de novo in PP-Rib-P synthetase-superactive cells appeared to reflect saturation of the rate-limiting amidophosphoribosyltransferase reaction for PP-Rib-P. In combination with accelerated purine excretion, increased channeling of newly formed purines into adenylates, and impaired conversion of AMP to IMP, fixed rates of purine synthesis de novo may condition cell lines with defects in HGPRT and PP-Rib-P synthetase to depletion of GTP with consequent growth retardation.

Aminopterin↗

PubMedDate not supplied

[Episodic disseminated inflammation of the central nervous system. Case mix review over a 13 year period].

INTRODUCTION: Episodic disseminated inflammation of the central nervous system (CNS) presents in processes that are difficult to differentiate, such as acute disseminated encephalomyelitis (ADE) and its multiphasic variants, and multiple sclerosis (MS). Magnetic resonance imaging allows these problems to be identified more frequently than in the past. PATIENTS AND METHODS: We carried out a retrospective study of the cases of episodic disseminated inflammation of the CNS, according to clinical features and compatible neuroimaging, at the Neuropaediatric Unit of the Hospital Infantil Miguel Servet, between May 1990 and August 2003. RESULTS: Of the 6777 children evaluated over this period, 10 met the eligibility criteria, with a minimum age at onset of 2 years and 2 months. In four cases there was a history of an infectious process or vaccination. Clinical involvement was multisymptomatic, the most frequent being ataxia, dysmetry, tremor, drowsiness, paresis and cranial nerve involvement. Six of them had cerebrospinal fluid disorders and only two presented disorders in the fundus oculi. Five of them were given corticoid treatment. Progress was favourable, except in two cases: one due to the persistence of a corticoid dependent optical neuropathy and the other because of dyskinesia in the right hand. DISCUSSION: Diagnosis of ADE is established by signs of multifocal clinical involvement and neuroimaging, and depends on a compatible progression. Prognosis is generally good and corticoids seem to be effective, at least in shortening the time the clinical features last. It is not possible to completely differentiate it from MS, especially in recurring forms. Clinical and magnetic resonance controls must be carried out.

Adolescent↗

PubMed2004

[Screening hepatitis B virus X-interactive proteins by yeast two-hybrid system].

BACKGROUND & OBJECTIVE: Hepatitis B virus-encoded X protein is a promiscuous transactivator and contributes to the development of hepatocellular carcinoma. Protein-protein interaction seems to be crucial for HBx transactivation. The aim of this study was to screen and identify the proteins which interact with hepatitis B virus (HBV) X protein by yeast two-hybrid system. METHODS: HBV X gene was amplified by polymerase chain reaction (PCR). HBV X bait plasmid, named pAS2-1-X, was constructed by yeast-two hybridization system 3 and verified by sequencing. pAS2-1-X was transformed into the yeast AH109, and X-BD fusion protein expressed in the yeast cells was confirmed by Western blot analysis. Yeast cells cotransformed with pAS2-1-X and normal human liver cDNA library were cultured in selective SC/-trp-leu-his-ade medium. The second screening was performed with beta-gal activity detection. The false positive clones were eliminated by segregation analysis and mating experiment. The real positive clones were amplified, sequenced, and analyzed with bioinformatics. RESULTS: Bait plasmid pAS2-1-X was successfully constructed. The result of Western blot analysis confirmed that pAS2-1-X correctly expressed X-BD fusion protein in the transformed yeast AH109. Ninety-seven clones grew in the selective SC/-trp-leu-his-ade medium; however, only one clone past through the beta-gal activity detection, segregation analysis, and mating experiment. The inserted cDNA fragment of positive clone showed high homology with Fis gene. CONCLUSION: Fis protein is a novel protein which can interact with X protein in vivo by yeast two-hybrid system.

Gene Library↗

PubMed2004

[Mutual interaction between hepatitis B virus antigen and metallothionein].

OBJECTIVE: To screen and identify the protein interacting with HBV antigen in hepatocytes. Then investigate the biological functions of hepatitis B virus antigen in the pathogenesis of hepatitis B and seek effective methods to prevent and treat it. METHODS: The yeast two-hybrid system-3 technique was used to construct HBV PreS2, HBeAg, HBcAg, HBxAg bait plasmids. The bait plasmids transformed the yeast AH109 and expressed themselves in it. After being identified by SDS-PAGE and Western blot, the AH109 yeast was mated with yeast Y187 containing liver cDNA library plasmid in 2 x YPDA medium to form diploid yeast and was then plated on synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) and synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) containing x-alpha-gal for screening. Plasmids of blue colonies were extracted and transformed into Escherichia coli, then analyzed by DNA sequencing and bioinformatics. To further prove the interaction between HBV antigen and metallothionein, translation was performed by using reticulocyte lysate and coimmunoprecipitation was displayed in vitro. RESULTS: Genes coding for HBV antigen binding protein were successfully cloned and metallothionein was found in that protein. The interaction between HBeAg, HBcAg and HBxAg and metallothionein were further proved by coimmunoprecipitation in vitro. CONCLUSION: The interaction between HBV antigen and metallothionein indicates that metallothionein may participate in the pathogenesis of hepatitis B

Hepatitis B↗

PubMed2004

[Study on binding of HBeAg to CD81].

AIM: To investigate the interaction between HBeAg and CD81. METHODS: The CD81 gene was amplified by RT-PCR from HepG2 cells. The recombinant expression vector pGADT7-CD81 was constructed by routine molecular biological method. The auxotroph yeast cells were cotransfected with pGADT7-CD81 and pGBKT7-eAg and plated on synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) containing x-alpha-gal. The binding of HBeAg to CD81 was also detected in vitro translation and coimmunoprecipitation test. RESULTS: The recombinant expression vector was constructed and confirmed by restriction enzyme (EcoR I and BamH I) digestion and DNA sequencing analysis. The positive yeast clones could grow and from blue colonies on SD/-Trp-Leu-His-Ade/x-alpha-gal medium. The result of immunocoprecipitation showed that HBeAg could bind to CD81. CONCLUSION: HBeAg can bind to CD81, suggesting that CD81 plays an important role in the pathogenesis of HBV.

Antigens, CD↗

PubMed2004

[Fourier transform infrared spectra analysis of nucleic acid in human breast tissue].

DNA molecular constitution is damaged to result in the inductive variation of base structure in female breast tissue under the influence of physical factors and chemical factors. The present research indicated that the FTIR spectra can reflect sensitively the change in the constitution. Our findings indicated that normal, benign and cancerous breast tissues are different in constitution and content of protein, nucleic acid and sugar, comparing the FTIR spectra, deconvolution spectra and date analysis. The result indicated that the content of collagen protein and nucleic acid increased obviously in cancerous tissue, while the content of glycoprotein increased gradually except mucinous carcinoma. After extracting nucleic acid of breast tissue, we further investigated the difference of constitution and content of base and phosphate by comparing normal, benign and cancerous breast tissues. The spectra and spectral data showed that the degree of hydrogen-bonding of base ring guanine (Gue) increased in cancerous tissue, base ring adenine (Ade) presented mostly in oxydic 8-OH-Ade via the attack of the *OH in cancerous breast, the peak position shifted to higher wave number with enhancing of C=N vibration, and the content of phosphate increased. After deconvolution, the ratio A1080/A1050 showed that the amount of PO2- relative to C-O increased from normal to cancerous breast tissues. It provided a important basis to study the mechanism of cancerization from molecular biology and molecular medicine.

Adenine↗

PubMed2005

[Screening of HBeAgTP interacting proteins in hepatocytes with yeast-two hybrid technique].

OBJECTIVE: To screen proteins in hepatocytes interacting with HBeAg transactivated protein (HBeAgTP) with yeast-two hybrid technique for investigating the biological functions of HBeAgTP. METHODS: Suppression subtractive hybridization (SSH) and bioinformatics techniques were used for screening and cloning of the target genes transactivated by HBeAg. The HBeAgTP gene was amplified by polymerase chain reaction (PCR) and HBeAgTP bait plasmid was constructed with yeast-two hybrid system 3, and then transformed into yeast AH109. The transformed yeast mated with yeast Y187 containing liver cDNA library plasmid in 2 x YPDA medium. Diploid yeast was plated on synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) and synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) containing X-gal for selecting two times and screening. After extracting and sequencing of plasmid from blue colonies, the results were analyzed by bioinformatics. RESULTS: HBeAgTP gene was successfully cloned and expressed in yeast cells. Fifteen genes in twenty-four positive colonies were obtained using yeast-two hybrid technique. CONCLUSION: HBeAgTP conjugated protein genes were successfully cloned, along with the genes involved in transcription and translation of proteins, immunoloregulation, materials and energy metabolism in vivo.

Hepatitis B e Antigens↗

PubMed2003

[Screening of new binding partners of CIKS with yeast two-hybrid system].

The NF-kappaB transcription factor plays important regulatory roles in inflammation, apoptosis, immune and stress responses. IkappaB kinase (IKK) composed of two catalytic subunits and a regulator subunit, acts as a key component of NF-kappaB activation pathway through phosphorylation of IkappaB, the inhibitor of NF-kappaB. CIKS (connection to IKK and SAPK), a newly identified cellular protein, is involved in NF-kappaB and JNK activation. Although it has been known that CIKS interacts with IKK complex, and activates both IKK and SAPK when overexpressed; the underling mechanisms are poorly understood. To better understand the physiological roles of CIKS, we have screened human HeLa MATCHMAKER cDNA library for new binding partners of CIKS by using the yeast two-hybrid system with truncated CIKS (151-574) as the bait. The yeast strain AH109 was sequentially transformed with the bait plasmid and the library. The transformants were screened on SD(-Leu/-Trp/-His/-Ade/ + X-alpha-gal)selective plates. Positive clones were restreaked on SD(-Leu/-Trp / + X-alpha-gal)plates three times to allow loss of some of the AD/library plasmids while maintaining selective pressure on both the DNA-BD and AD vectors. After 3 screenings on SD(-Leu/-Trp / + X-alpha-gal), the positive clones were further verified on SD(-Leu/-Trp/-His/-Ade/ + X-alpha-gal) plates. The inserts in AD/library plasmids were amplified by PCR and PCR products were characterized by Hae III digestion to eliminate the duplicates. After screening in selective plates, the positive AD/library plasmids were rescued via transformation of E. coli HB101 and the interactions of CIKS (151-574) with positive AD/library plasmids were further assessed by yeast two-hybrid analysis. Finally, the DNA sequences of the positive AD/library plasmids were determined and BLAST analysis against the databases was performed. The BLAST results indicate that the inserts in the positive plasmids encode RIKEN cDNA 473340F03, PLAC8, CD27BP (Siva-1), CDC5L, SnRNP smB, and DVL2. CDC5L is a key component of the multi-protein complex essential for the formation of pre-mRNA splicing product and is not required for spliceosome assembly. A role for CDC5L in the cell division cycle has been precious suggested as its overexpression of this protein in mammalian cells leads to a shortening G2 phase of the cell cycle, and a negative-dominant mutant of CDC5L lacking the N-terminal activation domain delays the G2 phase cell's entry into the mitosis. It has been reported that SnRNP smB participates in pre-mRNA splicing and CD27BP (Siva-1) binds to and inhibits BCL-XL-mediated protection against UV radiation-induced apoptosis and regulates T cell homeostasis. The functions of RIKEN cDNA 473340F03, PLAC8 and DVL2 are unknown. It has been suggested that CIKS functions as a critical component for cross-talk between NF-kappaB and JNK signaling pathways. IKK subunits, which have been demonstrated to interact with CIKS, were not shown up in this experiment. We speculate that the truncated CIKS (151-574) bait may not contain the binding domain that mediates the interaction of IKK subunits with CIKS. Taken together, the above results suggest that CIKS may play a role in cell regulation through interacting with various cellular proteins. Further investigations are required to characterize these interactions.

Adaptor Proteins, Signal Transducing↗

PubMed2005

Impact of newly available drugs on clinical progression in patients with virological failure after exposure to three classes of antiretrovirals.

OBJECTIVE: To study the prognosis of HIV-infected patients with virological failure after exposure to three classes of antiretroviral drugs (ARVs). DESIGN: Cohort study. SETTING: French Hospital Database on HIV. PATIENTS: Patients previously exposed to at least two nucleoside reverse transcriptase inhibitors (NRTIs), two protease inhibitors and one non-NRTI, with viral load (/L) values of > 5000 copies/ml after the exposure criteria were met and a new treatment initiated between 1998 and 2001 with VL > 5000 copies/ml. MAIN OUTCOME MEASURES: Risk of new AIDS-defining-events (ADEs) or death from first introduction of a drug never used before occurring between 1998 and 2001 defined as baseline. RESULTS: The main baseline characteristics of the 1092 patients were: previous ADE in 49% of cases, median CD4 cell count 181 microl, median VL 4.9 log10 copies/ml, median duration of ARV therapy 5.0 years and previous exposure to a median of nine ARVs. The crude progression rates were 20.1/100 patient-years among patients included in 1998, 15.1 in 1999, 11.1 in 2000 and 8.6 in 2001. After adjustment for baseline characteristics, the calendar year of inclusion was associated with the risk of clinical progression (P < 0.001). When the types of newly available drugs used at baseline or during follow-up were introduced into the model, year of inclusion was no longer associated with the risk of clinical progression (P = 0.42), while exposure to amprenavir/r, lopinavir/r, abacavir or tenofovir was associated with a lower risk. CONCLUSIONS: The clinical prognosis of heavily pretreated patients experiencing virological failure improved between 1998 and 2001, mainly thanks to the use of newly available drugs with more favourable resistance profiles.

Adult↗

PubMed2005

[In vitro activity of commercially available disinfectants against Stenotrophomonas maltophilia].

In vitro antibacterial efficacy of 14 commercially available disinfectants against the hospital pathogen Stenotrophomonas maltophilia was tested. The test disinfectants included 5 "pure" quartenary ammonium compounds (QACs) and 9 combination formulations with QACs as the major ingredient. Antibacterial efficacy was expressed as MIC, ED50 and inhibition of the rate of incorporation of radioactive precursors [14C] adenine and [14C] leucine. Based on their activity, the disinfectants were divided into four groups. Group I characterized by high inhibitory activity (MIC 0.045-0.09 microg/ml) comprised benzalconium chloride and Triquart. Group II included formulations with good antibacterial activity (MIC 0.19-0.78 microg/ml) while group III formulations showed MICs between 1.56 and 25 microg/ml. The less active Cetrimid (MIC 50-100 microg/ml) was classified into group IV. When effects on biosynthetic processes expressed as R values (IC50 Ade : IC50 Leu) were tested, R < 1 was only recorded for the following formulations: Hexaquart plus, Diesen forte, Sokrena, Forten, ID 212 and Cetrimid. Lower values of IC50 Ade and IC50 Leu are suggestive of effects on the synthesis of nucleic acids and proteins leading to inhibition of the two precursors. Endogenous respiration was almost 50% inhibited by Cetrimid and Microbac forte at a concentration of 0.78 microg/ml, and 100% inhibited by Almyrol, Diesen forte and Microbac forte at a concentration of 6.25 microg/ml and by FD312, Triquart, Hexaquart plus, Hexaquart S, ID 212, TPH 5225 at a concentration of 12.5 microg/ml.

Disinfectants↗

PubMed2005

[Screening and cloning of hepatitis C virus non-structural protein 4A interacting protein gene in hepatocytes].

OBJECTIVE: To investigate biological functions of hepatitis C virus (HCV) non-structural protein 4A (NS4A). METHODS: Yeast-two hybrid technique was performed to seek proteins in hepatocytes interacting with HCV NS4A. HCV NS4A bait plasmid was constructed by ligating the NS4A gene with carrier plasmid pGBKT7, then it was transformed into yeast AH109 (alpha type). The transformed yeast cells were amplified and mated with yeast cells Y187 (alpha type) containing liver cDNA library plasmid pACT2 in 2 x YPDA medium. Diploid yeast cells were plated on synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) and synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) containing X-alpha-gal for selection two times. After extracting plasmid from blue colonies, plasmid DNA was transformed into competent E.coli and analyzed by DNA sequencing and bioinformatics methods. RESULTS: Among twenty-two positive colonies there were eleven positive for metallothionein 2A, three for eukaryotic translation elongation factor 1 alpha 1, two for albumin, two for RNA binding motif protein 21, two for myomesin, one for cytochrome C oxidase II, and one for ATPase. CONCLUSIONS: Genes of HCV NS4A interacting proteins in hepatocytes were successfully cloned and the results pave the way for studying the biological functions of NS4A and associated proteins.

Carrier Proteins↗

PubMed2005

[Screening and cloning of hepatitis C virus non-structural protein 4B interacting protein gene in hepatocytes].

OBJECTIVE: To investigate biological functions of non-structural protein 4B (NS4B) of hepatitis C virus (HCV), yeast-two hybrid technique was performed to seek proteins in hepatocytes interacting with HCV NS4B. METHODS: HCV NS4B bait plasmid was constructed by ligating the NS4B gene with carrier plasmid pGBKT7 and transformed into yeast cells AH109 (type alpha). The transformed yeast cells were amplified and mated with yeast cells Y187 (alpha type) containing liver cDNA library plasmid pACT2 in 2 x YPDA medium. Diploid yeast cells were plated on synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) and synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) containing x-alpha-gal for selecting two times. After extracting plasmid from blue colonies, plasmid DNA was transformed into competent Escherichia coli and analysed by DNA sequencing and bioinformatics. RESULTS: Five genes in eight positive colonies were obtained. There were one NADH dehydrogenase subunit 3, one cytochrome c oxidase subunit III, one retinol binding protein 4, one reticulon 3-A (RTN3) and one fibrinogen gamma polypeptide (FGG). CONCLUSION: Genes of HCV NS4B interacting proteins in hepatocytes were successfully cloned and the results paved the way for studying the biological functions of NS4B and associated proteins.

Blotting, Western↗