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Biomedical subjects

F Libert

Publications and source records attributed to F Libert.

63 records · Page 4Linked to original sources
PubMed1989

Antibodies to human thyroid peroxidase in autoimmune thyroid disease: studies with a cloned recombinant complementary deoxyribonucleic acid epitope.

Previous studies carried out by screening a lambda gt11 human thyroid cDNA library with serum samples from selected patients with Hashimoto's thyroiditis and a polyclonal antibody to porcine thyroid peroxidase (TPO) confirmed, at the molecular level, that TPO is a major component of the thyroid microsomal antigen (M). That investigation led to the isolation of a clone (C2) which encodes an 85-amino acid segment of TPO and harbors a major epitope recognized by serum from several patients with autoimmune thyroid disease that contained anti-M autoantibodies (MAb). In this study, C2 antigen that was produced as a beta-galactosidase fusion protein was used to establish an enzyme-linked immunoabsorbent assay for the detection of anti-C2 autoantibodies (C2Ab). C2Ab then were assayed in 191 patients with different autoimmune and nonautoimmune thyroid disorders, and 50 patients with nonthyroidal autoimmune diseases. The results were compared with the titers of anti-TPO antibodies (TPOAb; as detected by monoclonal antibody-assisted RIA) and MAb (as detected by passive hemagglutination). Positive C2Ab was found in the serum of 85 of 136 (63%) patients whose serum contained TPOAb and/or MAb. A significant positive correlation was found between the levels of C2Ab and those of TPOAb (r = 0.76; P less than 0.001) or MAb (r = 0.69; P less than 0.001), which was independent of the type of underlying autoimmune thyroid disorder. Low levels of C2Ab also were found in 10 of 105 (9%) serum samples that did not contain TPOAb. Western blot analysis carried out on the latter samples showed that in 2 samples the apparent C2Ab reactivity was due to the presence of antibodies reacting with beta-galactosidase. In conclusion, we confirmed the validity of screening lambda gt11 cDNA human thyroid libraries to better characterize thyroid autoantigens and demonstrated the feasibility of using recombinant proteins to establish diagnostic assays for autoantibodies.

Antibodies↗
PubMed1989

Control of thyroperoxidase and thyroglobulin transcription by cAMP: evidence for distinct regulatory mechanisms.

The expression of the genes coding for thyroglobulin (TG), and thyroperoxidase (TPO), are regulated by TSH. These effects are mediated by cAMP as they are reproduced by forskolin. In vitro run-on transcription assays performed on nuclei isolated from dog thyrocytes in culture or from dog thyroid slices, indicate that the forskolin-induced transcriptional stimulation of TG and TPO genes are very different. For the TG gene, the kinetics of transcriptional activation vary according to the experimental model: it is rapid (1 h) in thyroid slices and slow (8 h) in primary cultures. In contrast, TPO induction is rapid in both cases. In primary cultures, insulin is responsible for the basal level and for a part of forskolin-induced TG transcription, whereas TPO transcription is not affected by insulin. The forskolin-induced increase of TG transcription requires ongoing protein synthesis, as it is blocked by cycloheximide. TPO gene transcription is unaffected by cycloheximide. Taken together with previous data on the two genes, our results suggest that while TPO regulation corresponds to the classical model of genes in which the promoter is regulated directly via cAMP regulatory elements, TG gene regulation involves the synthesis of an intermediary, rapid turnover trans-acting protein.

Animals↗
PubMed1988

Transcriptional regulation of the thyroperoxydase gene by thyrotropin and forskolin.

The expression of the gene coding for thyroperoxydase, the main enzyme involved in the synthesis of the thyroid hormones, is controlled by thyroid stimulating hormone. In vitro transcription assays performed on nuclei isolated from dog thyroid cells in primary culture showed that this control is at the transcription level, takes place rapidly (1 h) and is cyclic AMP-dependent as it is mimicked by forskolin. Insulin does not seem to be an important modulator of the thyroperoxydase gene expression.

Animals↗
PubMed1988

Regional localization of the gene for thyroid peroxidase to human chromosome 2pter----p12.

A 2.0-kb thyroid peroxidase cDNA of human origin was used as probe for Southern blot hybridization of genomic DNA from human somatic cells and human-rodent somatic cell hybrids. The results showed that the gene coding for human thyroid peroxidase is located on chromosome. 2. Further analysis of hybrids derived from Burkitt lymphoma cells carrying a (2;8)(p12;q24) translocation revealed that the gene maps to the region 2pter----p12.

Animals↗
PubMed1987

Thyroperoxidase, an auto-antigen with a mosaic structure made of nuclear and mitochondrial gene modules.

A lambda gt11 cDNA library was constructed from a normal human thyroid and screened with a rabbit anti-porcine thyroperoxidase antibody. A series of thyroperoxidase (TPO) clones were obtained which allowed determination of the complete primary structure of the protein. The library was also screened with serum from a patient with Hashimoto's thyroiditis, an autoimmune disease characterized by the presence in the serum of high titers of autoantibodies directed against the 'microsomal antigen' (McAg). Comparison of the cDNA sequences from TPO clones and McAg clones provides definite proof that the McAg is TPO. A short segment of TPO was characterized as bearing a major epitope involved in autoimmunity. The primary structure of TPO was 42% homologous to myeloperoxidase (MPO). It contains, in addition, a C-terminal extension with a membrane anchor region contiguous to two domains encoded by modules belonging to the EGF and C4b gene families. The existence in TPO of still another domain presenting a significant homology with a putative heme-binding region of cytochrome C oxidase polypeptide I raises the possibility that a mitochondrial gene module has contributed a piece to the evolution of a typical nuclear mosaic gene.

Amino Acid Sequence↗
PubMed1987

Preparation of single-stranded deoxyribonucleic acid probes using an immobilized template.

A new method for the easy preparation of specific single-stranded DNA fragments is presented. Recombinant M13 DNA containing the strand complementary to the sequence of interest is made partially double-stranded by elongating a conventional M13 sequencing primer. Following linearization by enzymatic digestion downstream from the insert (relative to priming site), this DNA is coupled to diazotized paper through its single-stranded (vector) portion. Subsequent denaturation of the double-stranded region generates an immobilized template strand. Successive runs of primed syntheses of the (desired) complementary strand can be realized using the same template. The copies are easily isolated by release upon denaturation. DNA probes prepared by this method have proven to be valuable tools for gene analysis.

DNA, Recombinant↗
PubMed1986

Methylation and expression of the human thyroglobulin gene.

The DNA methylation pattern at the 5'end of the human thyroglobulin gene has been determined in different tissues. Out of the four HpaII/MspI sites (5'-CCGG-3') present in this region, three were found to be non-methylated in thyroid DNA, while full methylation was observed in liver, salivary gland and sperm DNA. This demethylation therefore correlates with expression of the thyroglobulin gene. However, all four sites were found to be non-methylated in placental DNA, regardless of the activity of the gene.

Base Sequence↗
PubMedDate not supplied

Experience with living related liver transplantation in 63 children.

The incentive to develop intrafamilial living related liver transplantation (LRLT) originated from the shortage of cadaveric organ supply. We report our experience with LRLT in 63 children during 1993-1998 in the frame of a protocol approved by the Ethics Committee of our Institution. During this period, 152 potential intrafamilial (mostly parental) donors were evaluated; 44 (28.5%) were excluded because of surgical (n = 4), medical (n = 39) or psychosocial reason (n = 1). Out of 108 who matched all medical, surgical and psychological criteria of selection, 45 did not underwent living donation because their child received a cadaveric graft (n = 22; LRLT was their second option) or because one of the parents who had both been selected was chosen [by the surgical team because of more favourable anatomy (n = 8) or by mutual agreement between the two parents (n = 5)]. Sixty-three living donors (36 mothers, 24 fathers, one grand mother, one aunt and one uncle) underwent procurement of the left lobe (n = 52), the left lobe extended to part of segment IV (n = 8) or a left hepatectomy (n = 3) without mortality or any serious morbidity. Their median hospital stay was 7 days (range: 6-12); full physical rehabilitation and normalization of liver tests were usually obtained within three weeks. Their psychological follow-up did not disclose any longstanding serious sequellae. The median age of the recipients was 13 months (range 5-189); 30 were younger than one year at the time of transplant. Their median weight was 8.1 kg (range: 4.3 to 60); 36 had an actual weight under 10 kg. Fifty-two received an ABO identical and 11 received an ABO compatible transplant. The native liver diseases were similar to common data in children, with biliary atresia being the most frequent indication (74.6%). The median weight of the graft was 260 gr (range: 138-680) with a median ratio between the graft weight and the recipient body weight of 3.17% (range: 0.75-8.08). All grafts were implanted orthotopically with semi-microvascular reconstruction of the hepatic vein, portal vein and hepatic artery [end to end anastomosis in 58 (2 arteries were reconstructed in 7 patients) and interposition of an iliac arterial allograft from the infrarenal aorta in 5]. Base line immunosuppression consisted of a triple drug regimen including steroids, Azathioprine and either Cyclosporine-Sandimmun (n = 9), Cyclosporine Microemulsion formulation-Neoral (n = 13) or Tacrolimus-Prograft (n = 41). Biopsy-proved acute rejection was treated with intravenous bolus of steroids; steroid-resistant acute rejection was treated by a switch from Cyclosporine to Tacrolimus or addition of Mycophenolate-Mofetil (Cellcept) in Tacrolimus treated patients. Actuarial patient survival was 91.8% and 89.6% after LRLT at one and five years post-transplant, respectively, and 87.5% and 82.8% at one and five years, respectively, in 90 patients who received a cadaveric graft during the same interval. Actuarial graft survival was 91.8% and 84.1% after LRLT at one and five years, respectively, and 76.4% and 73.3% at one and five years, respectively, after cadaveric transplants. Vascular thrombosis was observed in 9.5% of the patients (arterial thrombosis: 1.6%; portal thrombosis: 7.9%) without graft loss. Biliary complications were observed in 26.9% (bile leak from cut surface in 3.1%, anastomotic stricture in 22.2% and intrahepatic stricture in 1.5%); two patients died from septic shock possibly related to uncompletely relieved anastomotic stricture; all other biliary complications were successfully treated either conservatively or surgically. The incidence of acute rejection was 90.9% in 22 patients with Cyclosporine-based immunosuppression; acute rejection was corticoresistant in 50%. It was 46.3% in 41 patients with Tacrolimus-based immunosuppression (64% with Prograft in capsules and 18.7% with Prograft in granules); no acute rejection was corticoresistant. (ABSTRACT TRUNCATED)

Adult↗