Nasal or intramuscular immunization of mice with influenza subunit antigen and the B subunit of Escherichia coli heat-labile toxin induces IgA- or IgG-mediated protective mucosal immunity.

Local mucosal IgA antibodies play a central role in protection of the respiratory tract against influenza virus infection. Therefore, new-generation influenza vaccines should aim at stimulating not only systemic, but also local antibody responses. Previously, we demonstrated that the recombinant B subunit of the Escherichia coli heat-labile toxin (LTB) is a potent adjuvant towards nasally administered influenza subunit antigen.

Characterization of antigen-presenting properties of tumour cells using virus-specific cytotoxic T lymphocytes.

Immunotherapy of tumours by induction of tumour-specific cytotoxic T-lymphocytes (CTLs) will only be effective for tumours with a functional antigen processing and presentation machinery. However, many tumours are known to down-regulate expression of major histocompatibility complex (MHC) class I molecules and/or to impair antigen processing. It is therefore desirable to evaluate the ability of a given tumour to present antigenic epitopes before developing an immunotherapy protocol.

Mucosal immunogenicity and adjuvant activity of the recombinant A subunit of the Escherichia coli heat-labile enterotoxin.

The Escherichia coli heat-labile enterotoxin (LT) is an exceptionally effective mucosal immunogen and mucosal immunoadjuvant towards coadministered antigens. Although, in general, the molecular basis of these properties is poorly understood, both the toxic ADP-ribosylation activity of the LTA subunit and the cellular toxin receptor, ganglioside, GM1-binding properties of the LTB-pentamer have been suggested to be involved. In recent studies we found that GM1-binding is not essential for the adjuvanticity of LT, suggesting an important role for the LTA subunit in immune stimulation.

Mucosal immunoadjuvant activity of recombinant Escherichia coli heat-labile enterotoxin and its B subunit: induction of systemic IgG and secretory IgA responses in mice by intranasal immunization with influenza virus surface antigen.

The Escherichia coli heat-labile enterotoxin (LT) is a very potent mucosal immunogen. LT also has strong adjuvant activity towards coadministered unrelated antigens and is therefore of potential interest for development of mucosal vaccines. However, despite the great demand for such mucosal vaccines, the use of LT holotoxin as an adjuvant is essentially precluded by its toxicity.

Role of GM1 binding in the mucosal immunogenicity and adjuvant activity of the Escherichia coli heat-labile enterotoxin and its B subunit.

Escherichia coli (E. coli) heat-labile toxin (LT) is a potent mucosal immunogen and immunoadjuvant towards co-administered antigens. LT is composed of one copy of the A subunit, which has ADP-ribosylation activity, and a homopentamer of B subunits, which has affinity for the toxin receptor, the ganglioside GM1.

The role of ADP-ribosylation and G(M1)-binding activity in the mucosal immunogenicity and adjuvanticity of the Escherichia coli heat-labile enterotoxin and Vibrio cholerae cholera toxin.

The mucosal route of vaccination has attracted a great deal of attention recently. Not only is mucosal application of vaccines, for example, orally or intranasally, particularly convenient, it also offers the possibility to induce locally produced and secreted S-IgA antibodies in addition to systemic IgG antibodies. These IgA antibodies are known to play a key role in protection against pathogens that invade the host through mucosal surfaces.

Mutational analysis of the role of ADP-ribosylation activity and GM1-binding activity in the adjuvant properties of the Escherichia coli heat-labile enterotoxin towards intranasally administered keyhole limpet hemocyanin.

The Escherichia coli heat-labile enterotoxin (LT) is known for its potent mucosal immunoadjuvant activity towards co-administered antigens. LT is composed of one A subunit, which has ADP-ribosylation activity, and a homopentameric B subunit, which has high affinity for the toxin receptor, ganglioside GM1. In previous studies, we have investigated the role of the LTA and LTB subunits in the adjuvanticity of LT towards influenza virus hemagglutinin (HA), administered intranasally to mice.

A new case of multiple mitochondrial enzyme deficiencies with decreased amount of heat shock protein 60.

Heat shock protein 60 (hsp60) is a mitochondrial matrix protein involved in the folding and correct assembly of polypeptides into complex mitochondrial enzymes. Its deficiency has recently been described as the most likely primary cause of congenital lactic acidaemia with multiple mitochondrial enzyme deficiencies in a female patient. We describe a new case of a girl with a substantially decreased amount of hsp60 in cultured fibroblasts.

Generation of heat shock protein-based vaccines by intracellular loading of gp96 with antigenic peptides.

Several studies have shown that immunization with heat shock proteins (HSPs) purified from tumors of virus-infected cells induces specific cytotoxic T-cell (CTL) activity. This immune response is directed against peptides bound to the HSPs rather than against the HSPs themselves. The peptides are derived from tumor- or virus-specific proteins which are degraded in the course of normal protein turnover and processing for presentation by MHC class I molecules.

Medium chain acyl-CoA dehydrogenase deficiency and fatal valproate toxicity.

A boy with delayed psychomotor development, attention deficit disorder, and therapy-resistant epilepsy was treated with valproate. The patient died of liver failure after 4 months of valproate treatment. Postmortem investigation of cultured fibroblasts suggested medium chain acyl-CoA dehydrogenase deficiency, an unexpected finding since the boy had not presented typical manifestations of this disease.

Mutants of the Escherichia coli heat-labile enterotoxin with reduced ADP-ribosylation activity or no activity retain the immunogenic properties of the native holotoxin.

The Escherichia coli heat-labile enterotoxin (LT) is a potent inducer of mucosal immune responses. In a previous study (L. DeHaan, W.

Transient expression of a mitochondrial precursor protein. A new approach to study mitochondrial protein import in cells of higher eukaryotes.

In order to study mitochondrial protein import in the context of whole cell metabolism, we have used the transfection technique based on Semliki Forest virus (SFV) to express a mitochondrial precursor protein within BHK21 cells and human fibroblasts. Recombinant SFV particles mediate a highly efficient, transient transfection of higher eukaryotic cells. The mitochondrial precursor protein used is a fusion protein consisting of the mitochondrial targeting sequence of Neurospora crassa ATPase subunit 9 and mouse dihydrofolate (H2folate) reductase.

Mucosal immunogenicity of the Escherichia coli heat-labile enterotoxin: role of the A subunit.

The Escherichia coli heat-labile enterotoxin (LT) is a potent mucosal immunogen, inducing high secretory as well as systemic antibody responses upon oral or intranasal (i.n.) administration.

Morphology of the mitochondria in heat shock protein 60 deficient fibroblasts from mitochondrial myopathy patients. Effects of stress conditions.

We have described two mitochondrial (mt) myopathy patients with reduced activities of various mt enzymes associated with significantly decreased amounts of heat shock protein 60 (hsp60). Experimental evidence suggested that the lack of hsp60 was the primary defect. Since hsp60 is essential for the proper folding of enzyme subunits in the mt matrix a partial deficiency of this protein can explain the observed defects of the mitochondria.

Decreased synthesis and inefficient mitochondrial import of hsp60 in a patient with a mitochondrial encephalomyopathy.

In a recent paper (Agsteribbe et al. (1993) Biochem. Biophys.

A fatal, systemic mitochondrial disease with decreased mitochondrial enzyme activities, abnormal ultrastructure of the mitochondria and deficiency of heat shock protein 60.

We report on a girl presenting with facial dysmorphic features and breathing difficulties upon birth. She was hypotonic, developed a metabolic acidosis, and died two days old of heart failure. Post-mortem examination revealed abnormalities of brain, lungs, heart and liver.

The human fumarylacetoacetase gene: characterisation of restriction fragment length polymorphisms and identification of haplotypes in tyrosinemia type 1 and pseudodeficiency.

Deficiency of human fumarylacetoacetase (FAH) activity results in hereditary tyrosinemia type I. Using the restriction enzymes BglII, KpnI and StuI and a 1.3-kb cDNA probe for the FAH gene, we have found 6 restriction fragment length polymorphisms (RFLPs).

Identification of three human pseudogenes for subunit VIb of cytochrome c oxidase: a molecular record of gene evolution.

Three pseudogenes for the nuclear-encoded subunit VIb of cytochrome c oxidase (COX) were isolated by screening a human genomic library with cloned human cDNA coding for COX subunit VIb. The nucleotide sequences of the pseudogenes, designated psi COX6b-1, psi COX6b-2 and psi COX6b-3, were determined. Pseudogene psi COX6b-1 bears all the hallmarks of a processed pseudogene and diverged from the parental gene after the divergence of man and cow.

Nucleotide sequence of the last exon of the gene for human cytochrome c oxidase subunit VIb and its flanking regions.

A human genomic clone encompassing the last exon of the gene for cytochrome c oxidase subunit VIb and a human genomic clone containing the most distal end of this gene were characterized. The last exon of the gene codes for the 17 C-terminal amino acid residues of the subunit and the 3' noncoding region. Downstream from the gene we found a single base difference between the DNA sequences of the two genomic clones.

Confocal scanning laser microscopy of mitochondria: a possible tool in the diagnosis of mitochondrial disorders.

This paper describes a non-invasive method for the study of mitochondrial morphology in cultured human skin fibroblasts by confocal scanning laser microscopy after staining the mitochondria with 2-[4-(dimethylaminostyryl]-1-methylpyridinium iodide. This method is applied to compare mitochondria in fibroblasts from healthy individuals and from patients with mitochondrial myopathy. In most cases there is a striking swelling of the patient's mitochondria and a loss of fine structure.

Isolation of cDNAs encoding subunit VIb of cytochrome c oxidase and steady-state levels of coxVIb mRNA in different tissues.

A full-length cDNA clone specifying the nuclear-encoded subunit VIb of human cytochrome c oxidase (COX) was isolated from a human skeletal muscle cDNA expression library. This was done with antiserum directed against the group of subunits VIa, b and c of bovine heart COX. A potential ribosome-binding site was located immediately upstream from the initiation codon.

Oxidative phosphorylation in human muscle in patients with ocular myopathy and after general anaesthesia.

The fuel preference of human muscle mitochondria has been given. Substrates which are oxidized with low velocity cannot be used to detect defects in oxidative phosphorylation. After general anaesthesia, the oxygen uptake with the different substrates is much lower than after local analgesia.

Duplication of the tRNA(MMet) and tRNA(Cys) genes and of fragments of a gene encoding a subunit of the NADH dehydrogenase complex in Neurospora grassa mitochondrial DNA.

Neurospora crassa mitochondrial DNA (mtDNA) contains duplications of the tRNA(MMet) gene upstream of a gene (ND2) encoding a subunit of the NADH dehydrogenase complex and of the tRNA(Cys) gene which is found downstream of the apocytochrome b gene. Both duplicated genes are located upstream of the small rRNA gene. The duplications are extended to flanking sequences.