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K Luo

Publications and source records attributed to K Luo.

At least 55 records · Page 3Linked to original sources
PubMed1999

Expression of a glycosylphosphatidylinositol-linked Manduca sexta aminopeptidase N in insect cells.

Aminopeptidase N (APN; EC 3.4.11.2) is an exopeptidase that is attached to cell membranes by a hydrophobic amino-terminal stalk in vertebrates or a glycosylphosphatidylinositol (GPI) anchor in insects. In this study, we report the cloning, expression, and characterization of an aminopeptidase N from Manduca sexta midgut. The full-length aminopeptidase N cDNA (APN1a) encodes a 995-amino-acid protein. The predicted amino acid sequence differs by 8 amino acids from M. sexta APN1. These different amino acids do not modify any putative glycosylation or glycosylphosphatidylinositol anchor sites. The full-length cDNA was cloned into an expression plasmid, pHSP-HR5, and transiently expressed in an insect cell line derived from Spodoptera frugiperda (Sf21 cells). Immunoblot analysis with anti-APN antiserum showed that APN1a expressed in Sf21 cells is the same size (120 kDa) as APN found in midgut brush border membranes. After treatment with phosphatidylinositol-specific phospholipase C (PIPLC), anti-cross-reacting determinant antibody specific for PIPLC cleavage products recognized the expressed 120-kDa APN1a, but not endogenous Sf21 proteins, indicating that APN1a has an intact glycosylphosphatidylinositol anchor. These results are evidence that Sf21 cells synthesize few, if any, endogenous GPI-linked proteins. Immunofluorescence staining showed that the expressed APN1a was located on the surface of Sf21 cells.

Amino Acid Sequence↗
PubMed1999

T1762/A1764 variants of the basal core promoter of hepatitis B virus; serological and clinical correlations in Chinese patients.

BACKGROUND: A double variant in the basal core promoter, converting nucleotide 1762 from A to T (T1762) and nucleotide 1764 from G to A (A[764), has been described in patients with chronic hepatitis B infection. Its prevalence and significance in Chinese chronic HBV carriers are unknown. METHODS: We studied 177 Chinese patients with chronic HBV infection (chronic hepatitis/asymptomatic: 89/88; hepatitis B e antigen positive/negative: 84/93). The double variant was detected by mismatched polymerase chain reaction and restriction fragment length polymorphism analysis. The reliability of this method was verified by sequencing in 41 serum samples with 100% matching. RESULTS: The double variant T1762/ A1764 was found in 52 of 89 patients with chronic hepatitis, but in only 6 of 59 asymptomatic carriers (p<0.001). The prevalence was significantly lower in hepatitis B e antigen positive patients (23/84) than in hepatitis B e antigen negative patients (35/64) (p<0.005). Precore variant, A1896 was detected in 40 individuals; 31 of them suffered from chronic hepatitis and 9 were asymptomatic (p<0.001). A combination of both variants T1762/A1764 and A1896 was seen in 3 of 59 asymptomatic and 22 of 89 patients with chronic hepatitis (p<0.005). CONCLUSIONS: Mismatched polymerase chain reaction with restriction fragment length polymorphism provides a reliable, easy and fast method for detection of the presence of the T1762/A1764 variant. In Chinese chronic hepatitis B carriers, T1762/A1764 variant was associated with both active liver disease and hepatitis B e antigen negativity.

Adult↗
PubMed1999

Toxicity, binding, and permeability analyses of four Bacillus thuringiensis Cry1 delta-endotoxins using brush border membrane vesicles of Spodoptera exigua and Spodoptera frugiperda.

The binding and pore formation properties of four Bacillus thuringiensis Cry1 toxins were analyzed by using brush border membrane vesicles from Spodoptera exigua and Spodoptera frugiperda, and the results were compared to the results of toxicity bioassays. Cry1Fa was highly toxic and Cry1Ac was nontoxic to S. exigua and S. frugiperda larvae, while Cry1Ca was highly toxic to S. exigua and weakly toxic to S. frugiperda. In contrast, Cry1Bb was active against S. frugiperda but only marginally active against S. exigua. Bioassays performed with iodinated Cry1Bb, Cry1Fa, and Cry1Ca showed that the effects of iodination on toxin activity were different. The toxicities of I-labeled Cry1Bb and Cry1Fa against Spodoptera species were significantly less than the toxicities of the unlabeled toxins, while Cry1Ca retained its insecticidal activity when it was labeled with 125I. Binding assays showed that iodination prevented Cry1Fa from binding to Spodoptera brush border membrane vesicles. 125I-labeled Cry1Ac, Cry1Bb, and Cry1Ca bound with high-affinities to brush border membrane vesicles from S. exigua and S. frugiperda. Competition binding experiments performed with heterologous toxins revealed two major binding sites. Cry1Ac and Cry1Fa have a common binding site, and Cry1Bb, Cry1C, and Cry1Fa have a second common binding site. No obvious relationship between dissociation of bound toxins from brush border membrane vesicles and toxicity was detected. Cry1 toxins were also tested for the ability to alter the permeability of membrane vesicles, as measured by a light scattering assay. Cry1 proteins toxic to Spodoptera larvae permeabilized brush border membrane vesicles, but the extent of permeabilization did not necessarily correlate with in vivo toxicity.

Journal Article↗
PubMed1999

[Cytotoxic effect of HepG2 cell expressing CD95L in an autocrine and paracrine way].

OBJECTIVE: The clearance of HBV-infected hepatocyte is mainly mediated by the CTL expressing CD95L. We have found that the hepatocytes also express CD95L in hepatitis B, a molecule with cytotoxic effect. This study was to elucidate its effect mechanism. METHODS: The HepG2 cell was induced to express CD95L and coincubated with HepG2.2.15 cell. The supernate from the induced HepG2 cell culture was added to another untreated HepG2 fraction. The apoptosis rates were examined with flow cytometry. The apoptosis of individual HepG2 cell incubation was also examined. RESULTS: The HepG2 cell expressing CD95L was able to kill the HepG2.2.15 cell with CD95 expression, apoptotic rates were 16.5% at 24 h and 43.0% at 48 h incubation. The supernate induced apoptosis rates of 38.7% and 73.3%, and the suicide rates of individual HepG2 cell incubation were 43.8% at 24 h. All these data were higher than those in controls. The apoptosis could be blocked with addition of anti-CD95L monoclonal antibody. CONCLUSION: The HepG2 cell expressing CD95L mediated cytotoxic effect by an autocrine and paracrine mechanism.

Apoptosis↗
PubMed1999

Hot spot mutations of hepatitis B virus pre-C/C gene and its promotor in Chinese patients and the clinical implications.

OBJECTIVE: To study if the functions of C gene and its promotor are related to the pathogenesis of hepatitis B and to describe their hot spot mutations in Chinese patients. DATA SOURCES: We have studied this subject in recent years. A mini-review is based on some unpublished works. DATA SELECTION: The sequence data of C gene and core promotor (cp) of hepatitis B virus (HBV) were analysed in the asymptomatic carriers (AsC) and the patients of hepatitis with various severity. RESULTS: Pre-C A1896 mutation occurred in 67% of anti-HBe-positive and 38% of HBeAg-positive cases. All were chronic infections. In C gene, 8 missense mutations in the segment of codon 48-60, and 28 in codon 84-101 were found, more in severe patients. The cp mutations of nt 1762 and nt 1764 occurred more in HBeAg-negative than in positive cases (49% vs 20%), and more in patients than in AsC (56% vs 10%). CONCLUSIONS: A1896 often occurred in nature infection course. The cp and clustering C gene mutations would more frequently lead to prolonged active infection and advanced liver diseases.

Base Sequence↗
PubMed1999

[Clearance of intrahepatic hepatitis B virus by a nonimmune mechanism].

OBJECTIVE: The clearance of intrahepatic hepatitis B virus (HBV) is mainly by the cytotoxicity of immune cells with ways including CD95 ligand/CD95 induction. We found that the liver cells do express CD95 ligand (CD95L), a cytotoxic cell surface molecule. This paper studied the mechanism and implications of the hepatocytes themselves to clear the virus. METHODS: The HepG2 cells with induced CD95L and the HepG2.2.15 cells with constitutive CD95 expression were coincubated. The apoptotic cells were examined under fluorescence microscope and with flow cytometry. RESULTS: In chronic hepatitis B intrahepatic expressions of CD95L and/or CD95 were detected at both mRNA and protein level. Methotrexate induced CD95L expression of HepG2 cell. The HepG2 cells with CD95L could kill the HepG2.2.15 cells with CD95 molecule with apoptotic rates of 16.5% at 24h and 43.0% at 48h. This apoptosis effect could be blocked by using anti-CD95L antibody. CONCLUSION: The hepatocyte might induce apoptosis mutually and clear the virus themselves with a nonimmune mechanism.

Apoptosis↗
PubMed1999

[Establishment of a method for classification of HBV genome and it's application].

OBJECTIVE: By using the techniques of PCR and restriction fragment length polymorphism (RFLP), we established a classification method for HBV genome based on viral S gene of asymptomatic HBV carriers (AsC). METHODS: By using this method we classified HBV DNA of AsC living in Guangzhou, Shenyang, Beijing and Chongqing. RESULTS: Among the AsC of Guangzhou, genotype B was 32.8%, genotype C 42.7%, mixed genotypes B and C 23.0%, others 1.6%; among the AsC of Chongqing, genotype B was 35.0%, genotype C 40.0%, mixed genotypes B and C 25.0%; among the AsC of Beijing, genotype B was 25.0%, genotype C 50.0%, mixed genotypes B and C 25.0%; among the AsC of Shenyang, genotype B was 11.1%, genotype C 88.9%. The prevalent HBV strains in China are genotype C and genotype B, and genotype C is the main genotype in west China. CONCLUSION: The established method for genotyping is simple and convenient. Using this method, we also classified HBV DNA of the patients with chronic hepatitis B and liver cirrhosis living in Guangzhou, and found that mixed genotypes B and C was in 50.0% of the patients detected, suggesting the mixed infection might lead to a severe damage of the liver tissue.

Carrier State↗
PubMed1998

Infection with HIV and hepatitis C virus among injecting drug users in a prevention setting: retrospective cohort study.

OBJECTIVES: To estimate the incidence of HIV and hepatitis C virus and risk factors for seroconversion among a cohort of injecting drug users. DESIGN: Retrospective cohort study. SETTING: Primary healthcare facility in central Sydney. SUBJECTS: Injecting drug users tested for HIV-1 antibody (n=1179) and antibodies to hepatitis C virus (n=1078) from February 1992 to October 1995. MAIN OUTCOME MEASURES: Incidence of HIV-1 and hepatitis C virus among seronegative subjects who injected drugs and underwent repeat testing. Demographic and behavioural risk factors for hepatitis seroconversion. RESULTS: Incidence of HIV-1 among 426 initially seronegative injecting drug users was 0.17/100 person years (two seroconversions) compared with an incidence of hepatitis C virus of 20.9/100 person years (31 seroconversions) among 152 injecting drug users initially negative for hepatitis C virus. Incidence of hepatitis C virus among injecting drug users aged less than 20 years was 75.6/100 person years. Independent risk factors for hepatitis C virus seroconversion were age less than 20 years and a history of imprisonment. CONCLUSIONS: In a setting where prevention measures have contributed to the maintenance of low prevalence and incidence of HIV-1, transmission of hepatitis C virus continues at extremely high levels, particularly among young injecting drug users.

Adult↗
PubMed1998

Transcription elongation factor P-TEFb mediates Tat activation of HIV-1 transcription at multiple stages.

Tat stimulates human immunodeficiency virus type 1 (HIV-1) transcription elongation through recognition of the transactivation response (TAR) RNA stem-loop structure at the 5' end of nascent viral transcripts. Recently, a human transcription elongation factor P-TEFb, consisting of CDK9 kinase, cyclin T and other associated factors, has been shown to interact with Tat to restore Tat activation in HeLa nuclear extract depleted of P-TEFb. Here, we report the purification of a P-TEFb complex fraction containing epitope-tagged wild-type CDK9 or kinase-inactive CDK9 and five tightly associated polypeptides. Only wild-type P-TEFb complex with an active CDK9 kinase was able to hyperphosphorylate the C-terminal domain of RNA polymerase II and mediate Tat transactivation in P-TEFb-depleted HeLa nuclear extract. Tat also stimulated transcription elongation by recruitment of the P-TEFb complex to the HIV-1 promoter through a Tat-TAR interaction. A possible mechanism for P-TEFb to become associated with polymerase elongation complexes and function as a general elongation factor was demonstrated by an interaction of P-TEFb with double-stranded RNA molecules through an 87 kDa subunit. Finally, P-TEFb was found to interact with and phosphorylate Tat-SF1, a Tat cofactor required for Tat transactivation. Our data indicate that the various subunits of the human P-TEFb complex may play distinct roles at multiple stages to mediate Tat activation of HIV-1 transcription elongation.

Base Sequence↗
PubMed1998

[Study on the physiological activities of the metabolites from Paecilomyces arovirens].

The metabolites of Paecilomyces arovirens exhibited insecticidal activities to many inscct pcsts, including cotton aphid, Aphis gossypii, hawthorn Ispidcr mitc, Tetranychus viennensis and larvae of imported cabbage worm, Pietis rapae. A comparison was made between the metabolites of P. arovirens and naphthyl acetic acid (NAA) in wheat (Triticum aestivum) and water melon (Cucumis sativus) bioassays on their physiological effects. Results demonstrated that some of the metabolites physiological activities of P. arovirens are same to that of NAA, some are different between them. It indicate that there is a new kind or many kinds of plant growth regulator in the metabolites and need further analysized. The results can offer reference for exploiting a new insecticid which not only can control insect pests, but also increase plant growth.

Cucumis sativus↗
PubMed1997

Risk of Kaposi's sarcoma and oroanal sexual contact.

After contradictory findings from a number of previous studies, behavioral risk factors for Kaposi's sarcoma were examined in a case-control study of 202 people diagnosed with acquired immunodeficiency syndrome (AIDS) in 1991-1993 in Sydney, Australia. Cases comprised 67 men who developed Kaposi's sarcoma at or after a diagnosis of acquired immunodeficiency syndrome, and controls were 135 people who did not have Kaposi's sarcoma at the time of diagnosis of acquired immunodeficiency syndrome or during follow-up until 1995. Men who developed Kaposi's sarcoma were more likely to report having a history of sexually transmissible diseases and having engaged more frequently than controls in a number of sexual practices with casual partners in the period before they became aware of their human immunodeficiency virus (HIV) infection. However, the only sexual practice reported significantly more often by cases at the 0.05 significance level was insertive oroanal contact with casual partners (odds ratio = 2.6, 95 percent confidence interval 1.3-5.3). This association was not present for insertive oroanal contact with regular partners or for insertive oroanal contact after subjects became aware of their HIV infection. The relation was present both in men who had Kaposi's sarcoma at the time of interview and in those who developed it later. The relation was not affected by adjustment for time of HIV infection and diagnosis or for other sexual practices. These results can be interpreted as supporting the hypothesis that Kaposi's sarcoma in people with HIV is caused by an infectious agent transmitted by oral contact with feces.

Acquired Immunodeficiency Syndrome↗
PubMed1997

Positive and negative regulation of type II TGF-beta receptor signal transduction by autophosphorylation on multiple serine residues.

The type II transforming growth factor-beta (TGF-beta) receptor Ser/Thr kinase (TbetaRII) is responsible for the initiation of multiple TGF-beta signaling pathways, and loss of its function is associated with many types of human cancer. Here we show that TbetaRII kinase is regulated intricately by autophosphorylation on at least three serine residues. Ser213, in the membrane-proximal segment outside the kinase domain, undergoes intra-molecular autophosphorylation which is essential for the activation of TbetaRII kinase activity, activation of TbetaRI and TGF-beta-induced growth inhibition. In contrast, phosphorylation of Ser409 and Ser416, located in a segment corresponding to the substrate recognition T-loop region in a three-dimensional structural model of protein kinases, is enhanced by receptor dimerization and can occur via an intermolecular mechanism. Phosphorylation of Ser409 is essential for TbetaRII kinase signaling, while phosphorylation of Ser416 inhibits receptor function. Mutation of Ser416 to alanine results in a hyperactive receptor that is better able than wild-type to induce TbetaRI activation and subsequent cell cycle arrest. Since on a single receptor either Ser409 or Ser416, but not both simultaneously, can become autophosphorylated, our results show that TbetaRII phosphorylation is regulated intricately and affects TGF-beta receptor signal transduction both positively and negatively.

Activin Receptors, Type I↗
PubMed1997

SIR2 and SIR4 interactions differ in core and extended telomeric heterochromatin in yeast.

Yeast core telomeric heterochromatin can silence adjacent genes and requires RAP1, SIR2, SIR3, and SIR4 and histones H3 and H4 for this telomere position effect. SIR3 overproduction can extend the silenced domain. We examine here the nature of these multiprotein complexes. SIR2 and SIR4 were immunoprecipitated from whole-cell extracts. In addition, using formaldehyde cross-linking we have mapped SIR2, SIR4, and RAP1 along telomeric chromatin before and after SIR3 overexpression. Our data demonstrate that SIR2 and SIR4 interact in a protein complex and that SIR2, SIR3, SIR4, and RAP1 map to the same sites along telomeric heterochromatin in wild-type cells. However, when overexpressed, SIR3 spreads along the chromosome and its interactions are dominant to those of SIR4 and especially SIR2, whose detection is decreased in extended heterochromatin. RAP1 binding at the core region is unaffected by SIR3 overproduction and RAP1 shows no evidence of spreading. Thus, we propose that the structure of core telomeric heterochromatin differs from that extended by SIR3.

Blotting, Western↗
PubMed1997

Binding of Bacillus thuringiensis Cry1Ac Toxin to Aminopeptidase in Susceptible and Resistant Diamondback Moths (Plutella xylostella).

Bacillus thuringiensis Cry1Ac toxin bound to a 120-kDa protein isolated from the brush border membranes of both susceptible and resistant larvae of Plutella xylostella, the diamondback moth. The 120-kDa protein was purified by Cry1Ac toxin affinity chromatography. Like Cry1Ac-binding aminopeptidase N (EC 3.4.11.2) from other insects, this protein was eluted from the affinity column with 200 mM N-acetylgalactosamine. The purified protein had aminopeptidase activity and bound Cry1Ac toxin on ligand blots. Purified aminopeptidase was recognized by antibodies to the cross-reacting determinant found on phosphatidylinositol-specific phospholipase C-solubilized proteins. The results show that the presence of Cry1Ac-binding aminopeptidase in the brush border membrane is not sufficient to confer susceptibility to Cry1Ac. Furthermore, the results do not support the hypothesis that resistance to Cry1Ac was caused by lack of a Cry1Ac-binding aminopeptidase.

Journal Article↗
PubMed1997

[The comparison of conventional open cholecystectomy, laparoscopic cholecystectomy and minor-incision cholecystectomy].

To study the clinical value of minor incision for cholecystectomy, compared conventional open cholecystectomy (300 patients, group A), alparoscopic cholecytectomy (300 patients, group B), and minor-incision cholecystectomy (300 patients, group C) with regard to duration (day) of operation, amount of intraoperative bleeding, complications, time (day) of gastrointestinal function recovery, intravenous infusion, and expense and time (day) of hospitalization. The results showed that the operating time, amount of bleeding, duration of intravenous infusion, time of gastrointestinal function recovery and hospitalization in group A were significantly different from those in group B and C (P < 0.05), but the incidence in group A was lower than that in group B. The operating time, amount of intraoperative bleeding, and the time of gastrointestinal function recovery and intravenous infusion in group B were similar to those in group C, but there was a high rate of intraoperative and postoperative complication in group B (1.66%). There was no complication in group C (P < 0.05). Comparison of the effects among group A, B and C showed that it was superior in group C to those in group A and B.

Adolescent↗
PubMed1997

[In vitro expression of wild type and precore mutant woodchuck hepatitis virus].

Hepatitis B virus (HBV) precore 1896 and 1898 G-->A mutations are regarded as hot spots to study. Such mutants have been considered associating with molecular and biological changes of HBV and causing chronic hepatitis and severe forms of hepatitis, including fulminant hepatitis. Using oligonucleotide-directed mutagenesis, we have conducted the same point mutations in the precore gene of an infectious clone of woodchuck hepatitis virus (WHV-8). Transfection of mutant WHV DNA, 1896 mutant (WHV-M1) or both 1896 and 1898 mutants (WHV-M2) resulted in WHV replication. Southern blot and Northern blot tests showed that wild type and mutants both had similar DNA replication and transcription. It was concluded that the precore gene of the WHV genome is not essential for virus replication.

Animals↗
PubMed1997

[Hot-point mutations of viral C gene during hepatitis B virus infection].

To investigate the relationship of the HBV C gene mutations with the disease progress, the hotpoint mutations of pre- C stop28 and C region L97 were examined with restrictio-fragment-length-polymorphism (RFLP) technique in 91 hepatitis patients with diverse clinical features and HBe status. Both mutations were almost not seen in acute hepatitis B and chronic asymptomatic virus carriers; rarely in chronic persistent hepatitis and frequently in chronic active hepatitis (CAH) and active liver cirrhosis (ALC), accounting for 80% and 78% in CAH and ALC respectively. The pre-C mutant was mixed with wild strain in 11 of 31 patients who were HBeAg-positive CAH and ALC; on the other hand, the wild strain also coexisted with variant in anti-HBe-positive cases. So did the L97 in both HBeAg- and anti- HBe-positive cases. Possibly, both the mutant and the wild strain are usually in a relatively growth and decline status. So, the hot-point mutations of HBV C gene were closely related with the disease activity.

Adult↗