Biomedical subjects

R Lin

Publications and source records attributed to R Lin.

At least 127 records · Page 7Linked to original sources

PubMed1995

Differential transcriptional activation in vitro by NF-kappa B/Rel proteins.

Distinct NF-kappa B subunit combinations contribute to the specificity of NF-kappa B-mediated transcriptional activation and to the induction of multiple cytokine genes including interferon-beta (IFN-beta). To evaluate the regulatory influence of different homo- and heterodimers, NF-kappa B subunits were analyzed for transcriptional activity in vitro using test templates containing two types of NF-kappa B recognition elements (the human immunodeficiency virus type 1 enhancer and the IFN-beta-positive regulatory domain-II (PRDII) as well as IFN-beta PRDIII-PRDI-PRDII linked to the -56 minimal promoter of rabbit beta-globin. Recombinant NF-kappa B subunits (p50, p65, c-Rel, p52, and I kappa B alpha) and interferon regulatory factor 1 were produced from either Escherichia coli or baculovirus expression systems. Transcriptional analysis in vitro demonstrated that 1) various dimeric complexes of NF-kappa B differentially stimulated transcription through the human immunodeficiency virus enhancer or PRDII up to 20-fold; 2) recombinant I kappa B alpha specifically inhibited NF-kappa B-dependent transcription in vitro; and 3) different NF-kappa B complexes and interferon regulatory factor 1 cooperated to stimulate transcription in vitro through the PRDIII-PRDI-PRDII virus-inducible regulatory domains of the IFN-beta promoter. These results demonstrate the role of NF-kappa B protein dimerization in differential transcriptional activation in vitro and emphasize the role of cooperativity between transcription factor families as an additional regulatory level to maintain transcriptional specificity.

Animals↗

PubMed1995

Interferon alfa-2b for chronic hepatitis C: effects of dose increment and duration of treatment on response rates. Results of the first multicentre Australian trial. Australia Hepatitis C Study Group.

Two hundred and thirty patients with histologically proven chronic hepatitis C were randomized to receive one of the following treatment protocols: (a) 3 million units of interferon alfa-2b thrice weekly for 6 months, (b) 5 million units thrice weekly for 6 months, or (c) 3 million units thrice weekly for 2 years. The short-term response to treatment was defined by normal alanine aminotransferase for at least 3 months and until the end of treatment, and was confirmed by loss of hepatitis C viraemia in 42 (91%) of 46 cases as determined by reverse transcription-polymerase chain reaction. Short-term response to interferon alfa-2b was independent of the incremental dose, being 64% for 5 million units and 58% for 3 million units. Long-term response to interferon alfa-2b was defined by continued normality of alanine aminotransferase levels for at least 6 months after treatment withdrawal. The long-term response rates among responders treated for 6 months and those treated for 2 years were 29% and 54%, respectively (p < 0.001). Among all 18 patients tested, serum HCV-RNA was negative at both 6 and 12 months of follow-up in all long-term responders, and none have subsequently relapsed. Improvement in hepatic necroinflammatory changes was confirmed by quantitative histology (Scheuer score) in responders at the end of interferon alfa-2b treatment. The changes were significantly greater among those who had been treated for 2 years compared with those treated for 6 months (p < 0.05 and p < 0.02, respectively, for portal and lobular inflammation scores). Several pretreatment characteristics could be correlated with a favourable response to interferon alfa-2b. Thus, absence of cirrhosis was associated with a short-term response of 75%, while only 42% of patients with cirrhosis had a short-term response (p < 0.001). The frequency of short-term response to interferon alfa-2b also differed according to mode of disease acquisition, being best for injecting drug use (71%), less favourable for blood transfusion (56%) and worst for sporadic cases (43%) (p < 0.01). This observed difference, however, was not independent of histology on multivariate analysis. In summary, a 5 million unit dose of interferon alfa-2b failed to improve the short-term or long-term response to interferon alfa-2b treatment, but prolongation of interferon alfa-2b treatment to 2 years resulted in substantially improved long-term response rate.

Adolescent↗

PubMed1995

Antipyrine clearance and response to interferon treatment in patients with chronic active hepatitis C.

To determine whether hepatic metabolic function affects the response to interferon treatment, we measured antipyrine clearance (APC) in 85 patients with chronic active hepatitis C and compared the results with treatment outcome. Among 55 patients who responded to interferon by normalization of alanine transaminase (ALT), median APC before treatment was 0.47 (range, 0.12 to 0.98; normal range, 0.34 to 1.02 mL/min/kg body wt), a value that was significantly greater than in 30 nonresponders (0.23; 0.08 to 0.67 mL/min/kg body wt, P < .001). APC was closely associated with response to interferon. The response rate among cases with values > 0.25 mL/min/kg body weight was 79%, the same as in cases without cirrhosis. Cases without cirrhosis and with APC of > 0.25 mL/min/kg body weight had an 85% chance of responding to interferon; this was unlikely a simple reflection of histological activity, because the correlation with Scheuer score was poor in this subgroup (r = -.31, P < .05). A second, independent group of 43 patients was used to test the predictive value of APC (using 0.25 mL/min/kg body wt as a cut-off) for response to interferon treatment. In this group, APC correctly predicted positive response to interferon in 75% of cases. APC was also used to measure the effects of treatment on hepatic metabolic function. Regardless of outcome, there was no change in APC at the end of a 6-month course of interferon treatment. Six months later, however, improvement in APC (14%; P < .05) was evident among responders but not in those who had failed to respond to interferon.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1995

Present status of the use of cytokines as adjuvants with vaccines to protect against infectious diseases.

Vaccine adjuvants are expected to play an important role in enhancing the immunogenicity of existing and new-generation vaccines against infectious diseases. In particular, adjuvants should direct the immune response in the most appropriate manner--furthering, for example, an expanded B-cell response, a cytotoxic T-cell response, or a T-helper 1 or 2 subset response. While some noncytokine adjuvants have exerted potent effects, their modes of action are most likely mediated by cytokines. Several cytokines have already been shown to be efficient adjuvants in animal models and/or in clinical trials. The mechanisms of cytokine function must be better understood and the techniques for the use of cytokines improved if the full potential of these substances as vaccine adjuvants is to be realized. When used to best advantage, such adjuvants enhance the immunity induced by viral, bacterial, and parasitic vaccines and thereby promote efficient protection or even cure.

Adjuvants, Immunologic↗

PubMed1995

Regulation of human immunodeficiency virus type 1 and cytokine gene expression in myeloid cells by NF-kappa B/Rel transcription factors.

CD4+ macrophages in tissues such as lung, skin, and lymph nodes, promyelocytic cells in bone marrow, and peripheral blood monocytes serve as important targets and reservoirs for human immunodeficiency virus type 1 (HIV-1) replication. HIV-1-infected myeloid cells are often diminished in their ability to participate in chemotaxis, phagocytosis, and intracellular killing. HIV-1 infection of myeloid cells can lead to the expression of surface receptors associated with cellular activation and/or differentiation that increase the responsiveness of these cells to cytokines secreted by neighboring cells as well as to bacteria or other pathogens. Enhancement of HIV-1 replication is related in part to increased DNA-binding activity of cellular transcription factors such as NF-kappa B. NF-kappa B binds to the HIV-1 enhancer region of the long terminal repeat and contributes to the inducibility of HIV-1 gene expression in response to multiple activating agents. Phosphorylation and degradation of the cytoplasmic inhibitor I kappa B alpha are crucial regulatory events in the activation of NF-kappa B DNA-binding activity. Both N- and C-terminal residues of I kappa B alpha are required for inducer-mediated degradation. Chronic HIV-1 infection of myeloid cells leads to constitutive NF-kappa B DNA-binding activity and provides an intranuclear environment capable of perpetuating HIV-1 replication. Increased intracellular stores of latent NF-kappa B may also result in rapid inducibility of NF-kappa B-dependent cytokine gene expression. In response to secondary pathogenic infections or antigenic challenge, cytokine gene expression is rapidly induced, enhanced, and sustained over prolonged periods in HIV-1-infected myeloid cells compared with uninfected cells. Elevated levels of several inflammatory cytokines have been detected in the sera of HIV-1-infected individuals. Secretion of myeloid cell-derived cytokines may both increase virus production and contribute to AIDS-associated disorders.

Base Sequence↗

PubMed1995

Leucine-responsive regulatory protein: a global regulator of gene expression in E. coli.

The leucine-responsive regulatory protein (Lrp) regulates transcription of the many genes of the Lrp regulon, repressing some and activating others, some in response to L-leucine and some independent of it. The physiology and molecular biology of the regulon in Escherichia coli are summarized here. However, the high degree of conservation of the protein suggests that it has an important role in all enterobacteria. We suggest that this role is not only as a transcriptional regulator but also as a determinant of chromosome structure.

Bacterial Proteins↗

PubMed1995

Retrovirus-mediated transfer of nuclear factor-kappa B subunit genes modulates I kappa B alpha and interferon beta expression.

Nuclear factor (NF)-kappa B proteins regulate the transcription of numerous genes involved in the immune response, transcription control, and viral pathogenesis. To examine the effect of ectopic expression of NF-kappa B proteins on DNA-binding activity and gene expression, individual NF-kappa B subunit genes were introduced into NIH 3T3 cells via retrovirus-mediated gene transfer. Expression of NF-kappa B subunits RelA (p65), NF-kappa B1 (p105), NF-kappa B2 (p100), and c-Rel increased the basal level of nuclear NF-kappa B DNA binding in NIH 3T3 cells, whereas expression of delta RelA (p65 delta) and NF-kappa B2 (p52) subunits did not affect basal level activity. Tumor necrosis factor-alpha treatment of the NF-kappa B-expressing cells stimulated the induced level of DNA-binding activity, reflecting interaction between endogenous murine and transfected human NF-kappa B proteins. Interestingly, expression of RelA (p65), c-Rel, NF-kappa B1 (p105), NF-kappa B2 (p100), and NF-kappa B2 (p52) subunits increased I kappa B alpha protein levels from 3- to 30-fold, indicating that one mechanism to compensate for the increased expression of NF-kappa B proto-oncogenes was to increase the synthesis and/or stability of the regulatory I kappa B alpha protein. In addition, overexpression of RelA (p65), c-Rel, NF-kappa B2 (p100), and NF-kappa B2 (p52) altered the induction kinetics of IFN-beta mRNA after Sendai virus infection, whereas overexpression of NF-kappa B1 (p105) dramatically decreased IFN-beta mRNA induction.

3T3 Cells↗

PubMed1994

Subcellular redistribution of HTLV-1 Tax protein by NF-kappa B/Rel transcription factors.

The viral oncogene Tax derived from human T cell leukemia virus type I (HTLV-I) is a positive transcriptional activator of HTLV-1 gene expression. Tax is also able to indirectly stimulate transcription of several growth regulatory genes by an indirect mechanism via association with host transcription factors. One of the cellular targets of the trans-activating effects of Tax is the NF-kappa B/Rel family transcription factors, pleiotropic regulators of immunoregulatory, cytokine, and viral gene expression. Recent studies demonstrated that specific subunits of NF-kappa B (NFKB2(p 100) and c-Rel) were overexpressed in HTLV-I-infected and Tax-expressing cells. Furthermore, Tax physically associated with NFKB2(p 100). Monospecific antibodies directed against individual NF-kappa B subunits were generated and used to investigate the consequences of the interactions between Tax and NF-kappa B in a cotransfection-immunofluorescence assay. These studies demonstrate: (1) distinct compartmentalization of NF-kappa B precursors and products, (2) differential induction of the endogenous I kappa B alpha protein by transfected NF-kappa B subunits, (3) subcellular relocalization of Tax to the cytoplasm or nucleus depending on the coexpressed NF-kappa B subunit, and (4) Tax interaction with the Rel homology domain region of NFKB2. These studies indicate that the transcription modulatory influence of HTLV-I Tax may be significantly influenced by cytoplasmic-nuclear partitioning associated with the NF-kappa B proteins.

Animals↗

PubMed1994

A Salmonella protein that is required for resistance to antimicrobial peptides and transport of potassium.

The ability of invading pathogens to proliferate within host tissues requires the capacity to resist the killing effects of a wide variety of host defense molecules. sap mutants of the facultative intracellular parasite Salmonella typhimurium exhibit hypersensitivity to antimicrobial peptides, cannot survive within macrophages in vitro and are attenuated for mouse virulence in vivo. We conducted a molecular genetic analysis of the sapG locus and showed that it encodes a product that is 99% identical to the NAD+ binding protein TrkA, a component of a low-affinity K+ uptake system in Escherichia coli. SapG exhibits similarity with other E. coli proteins implicated in K+ transport including KefC, a glutathione-regulated efflux protein, and Kch, a putative transporter similar to eukaryotic K+ channel proteins, sapG mutants were killed by the antimicrobial peptide protamine in the presence of both high and low K+, indicating that protamine hypersensitivity is not due to K+ starvation. Strains with mutations in sapG and either sapJ or the sapABCDF operon were as susceptible as sapG single mutants, suggesting that the proteins encoded by these loci participate in the same resistance pathway. SapG may modulate the activities of SapABCDF and SapJ to mediate the transport of peptides and potassium.

Amino Acid Sequence↗

PubMed1994

Mutational analysis of interferon (IFN) regulatory factors 1 and 2. Effects on the induction of IFN-beta gene expression.

Interferon (IFN) regulatory factor 1 (IRF-1) and IRF-2 are structurally similar but functionally distinct transcription factors that bind to the positive regulatory domains I and III (PRDI/III) within the human IFN-beta promoter. To begin structure-function analysis of IRF-1 and IRF-2, the regulatory potential of carboxyl-terminal deletion mutants was analyzed by co-transfection studies in human cells and was correlated with DNA binding capacity. Transcriptional repression by IRF-2 was contained within the first 125 amino-terminal amino acids and correlated directly with IRF-2 DNA binding; deletion to a protein of 100 amino acids resulted in loss of repression and IRF-2 DNA binding. Thus, the carboxyl terminus appears dispensible for trans-repression. Hybrid constructs which fuse the DNA binding domain of IRF-1 and IRF-2 to the trans-activation domain of NF-kappa B p65 were also generated; both IRF-1/p65 and IRF-2/p65 chimeras were strong transcriptional activators. IRF-2-mediated repression was also dominant over trans-activation by these fusion proteins. The trans-activation region of IRF-1 resides in the carboxyl terminus, primarily carboxyl-terminal to amino acid 250. Mutation of three potential casein kinase II phosphorylation sites within the IRF carboxyl terminus failed to identify an essential site that contributes to IRF-1 trans-activation potential.

Base Sequence↗

PubMed1994

Comparative efficacy of topical ciprofloxacin for treating Mycobacterium fortuitum and Mycobacterium chelonae keratitis in an animal model.

Mycobacterium fortuitum and M. chelonae are the two most common causes of nontuberculous mycobacterial keratitis, and they may be difficult to differentiate at diagnosis. Mycobacterium fortuitum is generally more sensitive to ciprofloxacin in vitro than is M. chelonae. Using a rabbit model, we compared the efficacy of topical ciprofloxacin (3 mg/ml) against M. chelonae keratitis to its efficacy against M. fortuitum keratitis. After four days of therapy, ciprofloxacin significantly reduced the number of both organisms in treated eyes compared to untreated control eyes (both P values < .001). Mean culture ratios (colony-forming units in treated eye divided by colony-forming units in untreated eye for each rabbit) were used to compare efficacy between groups. When all treated animals were considered, no significant difference was found between groups (P = .13). When outlier values were excluded, ciprofloxacin was more effective against M. fortuitum than M. chelonae (P = .01). When treated and untreated eyes were compared after therapy in the M. fortuitum group, ciprofloxacin treatment was associated with a reduction in mean stromal infiltrate area (P = .03) and in the tendency to form satellite lesions (P = .07). A clinical effect was not observed in the M. chelonae group. Although ciprofloxacin is effective against both organisms, it appears to be less effective against M. chelonae than M. fortuitum in vivo.

Administration, Topical↗

PubMed1994

Thioallyl compounds: potent inhibitors of cell proliferation.

S-Allylmercaptocysteine was shown to inhibit vascular smooth-muscle and umbilical endothelial cell proliferation. Inhibition was dose-dependent and affected smooth-muscle cells more than endothelial cells. S-Allylmercaptocysteine was two orders of magnitude more potent than S-allylcysteine and cells grown in its presence showed distinct changes in their phosphorylation compared to untreated controls. Among the proteins whose phosphorylation was altered were GTP-activating protein, protein tyrosine phosphatase-1B and p34cdc2. We conclude that thioallyl compounds, natural constituents of garlic and known to inhibit malignant cells, can also reduce the proliferation of normal cells.

Animals↗

PubMed1994

Potential role of granulocyte-macrophage colony-stimulating factor as vaccine adjuvant.

The uses of GM-CSF as an immunomodulator and vaccine adjuvant are reviewed. GM-CSF has a variety of effects on immune responses: it induces class II major histocompatibility complex antigen expression on the surface of macrophages; it enhances dendritic cell maturation and migration; it results in a localized inflammation at the injection site; and it has marked effects on maturation of haematopoietic progenitor cells in the bone marrow. Animal and human studies suggest that administration of GM-CSF can increase antibody titres to foreign antigens. Monkeys injected with human interleukin (IL)-3 plus GM-CSF, at a different injection site, developed peak antibody titres which were 8- to 30-fold higher than those in monkeys injected with IL-3 alone. In a study of ovarian cancer patients receiving GM-CSF to prevent chemotherapy-induced neutropenia, two patients who had demonstrated a low titre of antithyroid antibodies prior to the study showed an increase in antibody titre and transient thyroiditis after administration of GM-CSF. Recently a GM-CSF/antigen fusion protein has been tested. An antibody corresponding to a specific idiotype expressed on B-cell lymphomas was fused to GM-CSF and injected into mice with B-cell lymphoma xenografts. The mice developed antibodies to the lymphoma and there was a protective effect against disease progression. Preliminary results of clinical trials using GM-CSF in humans suggest that it enhances antibody responses to hepatitis B vaccine. On the basis of these preliminary results, several clinical trials are being planned and it would appear that GM-CSF has potential as a vaccine adjuvant.

Adjuvants, Immunologic↗

PubMed1994

Demonstration of differences in vascular permeability in experimental tumors by use of 19F magnetic resonance imaging.

RATIONALE AND OBJECTIVES: In vivo assessment of tumor vascular permeability may provide useful information for chemotherapy treatment planning or for the assessment of treatment effectiveness. We aimed to assess vascular permeability in two tumor sublines as well as changes in vascular permeability with tumor growth by using 19F magnetic resonance imaging. METHODS: An emulsion of perfluorotributylamine was used as a tumor extravascular contrast agent for 19F MRI. The amount of emulsion that leaked into tumor interstitial space was analyzed qualitatively with imaging. A quantitative study of vascular permeability was done with a separate group of tumors by use of Evans blue dye. RESULTS: One tumor type was more permeable to both perfluorotributylamine emulsion and Evans blue than was the second tumor type. The difference was attributed to a difference in surface area for exchange. In larger tumors of both types, pooling of large amounts of perfluorocarbon occurred and was assumed to be attributable to hemorrhage or blood flow stasis or both. CONCLUSION: 19F MRI is capable of demonstrating the permeability of tumor vessels to macromolecular substances.

Adenocarcinoma↗

PubMed1994

Viral induction of the human beta interferon promoter: modulation of transcription by NF-kappa B/rel proteins and interferon regulatory factors.

Multiple regulatory domains within the -100 region of the beta interferon (IFN-beta) promoter control the inducible response of the IFN gene to virus infection. In this study, we demonstrate that the formation of NF-kappa B-specific complexes on the positive regulatory domain II (PRDII) precedes the onset of detectable IFN-beta transcription in Sendai virus-infected cells. By using NF-kappa B subunit-specific antibodies, a temporal shift in the composition of NF-kappa B subunits in association with the PRDII domain is detected as a function of time after virus infection. Furthermore, a virus-induced degradation of I kappa B alpha (MAD3) protein is observed between 2 and 8 h after infection; at later times, de novo synthesis of I kappa B alpha restores I kappa B alpha to levels found in uninduced cells and correlates with the down regulation of IFN-beta transcription. In cotransfection experiments using various NF-kappa B subunit expression plasmids and two copies of PRDII/NF-kappa B linked to a chloramphenicol acetyltransferase reporter gene, we demonstrate that expression of p65, c-Rel, or p50 or combinations of p50-p65 and p65-c-Rel differentially stimulated PRDII-dependent transcription. Coexpression of I kappa B alpha completely abrogated p65-, c-Rel-, or p65-p50-induced gene activity. When the entire IFN-beta promoter (-281 to +19) was used in coexpression studies, synergistic stimulation of IFN-beta promoter activity was obtained when NF-kappa B subunits were coexpressed together with the IFN regulatory factor 1 (IRF-1) transcription factor. Overexpression of either I kappa B or the IRF-2 repressor was able to abrogate inducibility of the IFN-beta promoter. Thus, multiple regulatory events--including differential activation of DNA-binding NF-kappa B heterodimers, degradation of I kappa B alpha, synergistic interaction between IRF-1 and NF-kappa B, and decreased repression by I kappa B and IRF-2--are all required for the transcriptional activation of the IFN-beta promoter.

Animals↗

PubMed1994

[Effect of indirect fluorescent antibody test for the diagnosis of amoebiasis].

Indirect fluorescent antibody test (IFAT) was performed by the method of [symbol: see text] (1971) with some slight modifications for the diagnosis of amoebiasis. The antigen used was prepared from 48-hour culture of Entamoeba histolytica (strains G2, G3, and G5) in LAS diphasic medium and only those amoebae clinging the culture were collected. The serum samples or blood drops on filter paper were positive by the improved IFAT in all of the 54 cases of amoebic liver abscess and the positive reactions of most cases were very marked. Of 23 cases of acute amoebic dysentery, 19 were positive, the positive rate being 82.6%. The results also show that 42 cases of other diseases and 40 healthy persons were all negative.

Animals↗

PubMed1993

Comparison of topical antibiotics for treating Mycobacterium fortuitum keratitis in an animal model.

The efficacy of three topical antibiotic treatments for Mycobacterium fortuitum (strain ATCC-6841) keratitis were compared in rabbits. Rabbits were treated with ciprofloxacin (3 mg/ml) or clarithromycin (20 mg/ml) or a combination of amikacin (100 mg/ml) and vancomycin (50 mg/ml). All three treatments significantly reduced the number of organisms in treated eyes compared to untreated, control eyes (all P values < .001). No significant difference in treatment efficacy was found between the three treatment groups (all P values > or = .48), although ciprofloxacin (3 mg/ml) was more effective than clarithromycin (20 mg/ml) after excluding outliers (P = .01). All treatments stabilized or reduced the size of stromal infiltrates after four days of therapy, whereas infiltrates continued to enlarge in untreated eyes. These results suggest that topical clarithromycin, topical ciprofloxacin, and combined amikacin and vancomycin may all be clinically useful for treating M. fortuitum keratitis. Both clarithromycin and ciprofloxacin were better tolerated than combined amikacin and vancomycin. This study supports the further development of clarithromycin, a new macrolide antibiotic, as a topical drug for treatment of M. fortuitum keratitis.

Administration, Topical↗

PubMed1993

Molecular, functional, and evolutionary analysis of sequences specific to Salmonella.

In that salmonellae have been implicated in an unprecedented array of diseases, sequences found to be specific to this species are often thought to be involved in the virulence attributes not seen in other enteric bacteria. To identify the molecular, genetic, and phenotypic characteristics that differentiate bacterial species, we analyzed five cloned DNA fragments that were originally described as being confined to Salmonella. Most of these segments mapped to unique positions on the Salmonella typhimurium chromosome indicative of independent evolutionary events, and three had G+C contents considerably lower than that of the Salmonella genome, suggesting that they arose through horizontal transfer. The nucleotide sequence was determined for one of the clones exhibiting an atypical base composition. This 4.9-kb fragment contained an open reading frame with structural similarity to the LysR family of transcriptional regulators. Strains harboring deletions in this region were tested for > 120 phenotypic characteristics including the effects on a collection of environmentally regulated lac gene fusions. In addition, all deletion strains behaved like the wild-type parent when tested for virulence in mice.

Amino Acid Sequence↗