Biomedical subjects
Publications and source records attributed to M Renedo.
Motivated by path-integral numerical solutions of diffusion processes, PATHINT, we present a tree algorithm, PATHTREE, which permits extremely fast accurate computation of probability distributions of a large class of general nonlinear diffusion processes.
T cell non-Hodgkin's lymphomas are a heterogeneous group of lymphomas with poor prognosis, and whose genetic alterations are not well understood. Comparative genomic hybridization (CGH) is a technique that allows the identification of DNA imbalances without cytogenetic studies. We have studied 37 samples from 29 T cell non-Hodgkin's lymphomas (25 peripheral and four lymphoblastic lymphomas) by CGH in order to detect DNA sequence copy number changes of putative importance in the biology and prognosis of these neoplasms. We detected abnormal CGH profiles in 16/27 (59%) of samples at diagnosis, a ratio that increased to 66% (23/37) when we included the relapsed samples. The most common recurrent changes were gains related to the X chromosome, either the whole chromosome or partially the Xq26-27 bands (19%). Other recurrent changes included gains of bands 9q34, gains of chromosomes 17, 19, and 20, and complete or partial deletions of chromosome 13 (10%). Cancer-related genes located at Xq26-28 region were analyzed by Southern blot and fluorescence in situ hybridization (FISH). Low level amplification of some of these genes was detected by this technique confirming the results obtained by CGH in this region. The detection of abnormal CGH profiles in these T cell lymphomas could have clinical implications. Patients with abnormal CGH profiles showed significant associations with advanced stage of disease, overexpression of P53, and higher proliferative index.
We developed a sequence-ready physical map of a part of human chromosome 12p12.3-p13.2 where the natural killer gene complex (NKC) is located. The NKC includes a cluster of genes with structure similar to that of the Ca(2+)-dependent lectin superfamily of glycoproteins that are expressed on the surface of most natural killer (NK) cells and a subset of T cells. These killer cell lectin-like receptors (KLR) are involved in NK target cell recognition, leading to activation or inhibition of NK cell function. We used a number of sequence-tagged site (STS) markers from this region to screen two large insert bacterial artificial chromosome (BAC) libraries and a bacteriophage P1-derived (PAC) chromosome library. The clones were assembled into contiguous sets by STS content analysis. The 72-BAC and 11-PAC contig covers nearly 2 Mb of DNA and provides an average marker resolution of 26 kb. We have precisely localized 17 genes, 5 expressed sequence tags, and 49 STSs within this contig. Of this total number of STS, 30 are newly developed by clone-end sequencing. We established the order of the genes as tel-M6PR-MAFAL (HGMW-approved symbol KLRG1)-A2M-PZP-A2MP-NKRP1A (HGMW-approved symbol KLRB1)-CD69-AICL (HGMW-approved symbol CLECSF2)-KLRF1-OLR1-CD94 (HGMW-approved symbol KLRD1)-NKG2D (HGMW-approved symbol D12S2489E)-PGFL-NKG2F (HGMW-approved symbol KLRC4)-NKG2E (HGMW-approved symbol KLRC3)-NKG2A (HGMW-approved symbol KLRC1)-LY49L (HGMW-approved symbol KLRA1)-cen. This map would facilitate the cloning of new KLR genes and the complete sequencing of this region.
A high protein tyrosine phosphatase (PTPase) activity is required to maintain circulating T lymphocytes in a resting phenotype, and to limit the initiation of T cell activation. We report that 15 of the currently known 24 intracellular PTPases are expressed in T cells, namely HePTP, TCPTP, SHP1, SHP2, PEP, PTP-PEST, PTP-MEG2, PTEN, PTPH1, PTP-MEG1, PTP36, PTP-BAS, LMPTP, PRL-1 and OV-1. Most were found in the cytosol and many were enriched at the plasma membrane. Only TCPTP and PTP-MEG2 had subcellular localizations that essentially excludes them from a direct role in early T cell antigen receptor signaling events. Overexpression of 6 of the PTPases reduced IL-2 gene activation, 3 of them thereby identified as novel candidates for negative regulators of TCR signaling. Our findings expand the repertoire of PTPases that should be considered for a regulatory role in T cell activation.
The human NK gene complex localized on chromosome 12p12.3 - p13.2 codes for several lectin-like receptor genes expressed by NK cells as well as by other hematopoietic cells. In this study, by using the expressed sequence tag database we identified a novel receptor gene, designated as killer cell lectin-like receptor, subfamily F, member 1 (KLRF1), encoding a putative type II transmembrane glycoprotein. The KLRF1 gene has been localized on the high-resolution physical map of chromosome 12p. The genomic structure of the KLRF1 gene and the existence of one spliced variant are also described. KLRF1 was expressed at the mRNA level in peripheral blood leukocytes, activated NK cells, monocytes and NK and myeloid cell lines. The presence of two immunoreceptor tyrosine-based inhibitory-like motifs within the cytoplasmic tail of KLRF1 suggests an inhibitory role in NK cell and monocyte activity.
THP-1 monocytic/macrophage cells were stimulated via their FcgammaR receptors with insoluble aggregates of human IgG and the production of the C-C chemokine monocyte chemoattractant protein (MCP)-1 assayed. A dose- and time-dependent production of MCP-1 comparable to that produced by the most potent agonists could be detected in the culture medium by a sensitive ELISA assay. This was accompanied by a parallel activation of the transcription factor NF-kappaB as judged from both the appearance of kappaB-binding activity containing p50/p65 NF-kappaB/Rel complexes in the nuclear extract and the disappearance of the NF-kappaB inhibitor IkappaB-alpha in the cell lysate. In contrast, IkappaB-beta and IkappaB-epsilon expression was not modified, thus pointing to the occurrence of a selective degradation of IkappaB-alpha under those conditions. Attempts to modulate MCP-1 production with compounds that display inhibitory effects on the activation of NF-kappaB such as the proteasome inhibitor N-acetyl-leucinyl-leucinyl-norleucinal, the antioxidant pyrrolidine dithiocarbamate and the salicylate derivative 2-hydroxy-4-trifluoromethylbenzoic acid showed a parallel effect on both MCP-1 production and NF-kappaB activation, thus pointing to the involvement of kappaB-binding sites on the transcriptional regulation of MCP-1 production. Our findings suggest the existence in monocytic cells of a signaling mechanism initiated by cross-linking of low-affinity FcgammaR, most likely of the FcgammaRII family since THP-1 cells do not express FcgammaRIII receptors, that involves activation of NF-kappaB associated to the proteolytic degradation of IkappaB-alpha and leads to the transcriptional up-regulation of MCP-1.
BACKGROUND: The Prader-Willi syndrome (PWS) is a neurogenetic disorder associated with abnormalities in the chromosomal region 15q11-13 of paternal origin. Most cases (65-85%) have a deletion involving the paternally derived chromosome and the remainder (20-25%) have a maternal uniparental disomy. Some patients have a defect in the imprinting process. We report the results of molecular, cytogenetic and clinical studies on 10 PWS patients. PATIENTS AND METHODS: 18 suspected patients were classified as PWS typical or not typical as they fulfilled or not the clinical criteria for PWS. Cytogenic studies-high resolution chromosome banding analyses (HRGTG) and fluorescence in situ hybridization (FISH) -and molecular chromosome genetic analyses--microsatellite markers and Southern blotting--were carried out from peripheral blood lymphocytes. RESULTS: PWS was confirmed in 10 probands. 8 fulfilled the clinical criteria for PWS and showed cytogenetic and/or molecular abnormalities. In 2 patients without clinical or cytogenetic data, diagnosis was confirmed by molecular methods only. Cytogenetic and molecular findings describe a characteristic clinical picture of PWS. CONCLUSIONS: Cytogenetic techniques (FISH and HRGTG) confirmed PWS diagnosis in 40% of cases, microsatellite studies in 70% of them and Southern blotting (the metilation test) in 100% cases. Southern blotting is the method of choice for rapid diagnostic testing of patients suspected of having PWS.
Natural killer (NK) cells preferentially express several type II glycoproteins of the calcium-dependent lectin superfamily. The genes coding for these molecules are clustered on the distal mouse chromosome 6 and on the rat chromosome 4 in a region designated the NK gene complex. To date, no definite evidence of the presence of a NK gene complex has been found in humans. Here we report the assignment by fluorescence in situ hybridization of the CD94 gene to human chromosome 12p12-p13, in the same region where the CD69 and NKG2A genes had been previously mapped. In addition, using a yeast artificial chromosome contig spanning this region we determined that the human CD94, NKG2A, NKG2C, NKG2E, and NKR-P1A (NKR) genes map to the short arm of chromosome 12. The distal to proximal position of these loci are: NKR- CD69 - CD94/NKG2A/NKG2C/NKG2E. These data demonstrate the existence of a human NK gene complex located within a 5.6 cM interval flanked by the genetic markers D12S397 and D12S89. The physical distance spanned by the NK gene complex in humans ranges between 0.7 and 2.4 megabases.
We present cytogenetic, immunologic, and molecular data obtained over 7 years from a family with ataxia telangiectasia (AT) including 2 affected children and their unaffected sibling, and their obligate heterozygous parents. In a period of 3 years, the T lymphocytes from both AT patients showed clonal rearrangements of chromosomes 7 and 14 at specific bands (7p13, 7q35, 14q12, and 14q32), where loci for the Ig and TCH genes are located. A molecular study was carried out on peripheral blood and bone marrow samples from both patients using Southern blot and PCR for Ig and TCR genes. A monoclonal population for TCR gamma was observed in one of the two affected children, but only in peripheral blood.
Alkaptonuria (AKU) occupies a unique place in the history of human genetics because it was the first disease to be interpreted as a mendelian recessive trait by Garrod in 1902. Alkaptonuria is a rare metabolic disorder resulting from loss of homogentisate 1,2 dioxygenase (HGO) activity. Affected individuals accumulate large quantities of homogentisic acid, an intermediary product of the catabolism of tyrosine and phenylalanine, which darkens the urine and deposits in connective tissues causing a debilitating arthritis. Here we report the cloning of the human HGO gene and establish that it is the AKU gene. We show that HGO maps to the same location described for AKU, illustrate that HGO harbours missense mutations that cosegregate with the disease, and provide biochemical evidence that at least one of these missense mutations is a loss-of-function mutation.
We analyzed 61 Angelman syndrome (AS) patients by cytogenetic and molecular techniques. On the basis of molecular findings, the patients were classified into the following 4 groups: familial cases without deletion, familial cases with submicroscopic deletion, sporadic cases with deletion, and sporadic cases without deletion. Among 53 sporadic cases, 37 (70%) had molecular deletion, which commonly extended from D15S9 to D15S12, although not all deletions were identical. Of 8 familial cases, 3 sibs from one family had a molecular deletion involving only 2 loci, D15S10 and GABRB3, which define the critical region for AS phenotypes. The parental origin of deletion, both in sporadic and familial cases, was exclusively maternal and consistent with a genomic imprinting hypothesis. Among sporadic and familial cases without deletion, no uniparental disomy was found and most of them were shown to inherit chromosomes 15 from both parents (biparental inheritance). A discrepancy between cytogenetic and molecular deletion was observed in 14 (26%) of 53 patients in whom cytogenetic analysis could be performed. Ten (43%) of 23 patients with a normal karyotype showed a molecular deletion, and 4 (13%) of 30 patients with cytogenetic deletion, del(15) (q11q13), showed no molecular deletion. Most clinical manifestations, including neurological signs and facial characteristics, were not distinct in each group except for hypopigmentation of skin or hair. Familial cases with submicroscopic deletion were not associated with hypopigmentation. These findings suggested that a gene for hypopigmentation is located outside the critical region of AS and is not imprinted.
The use of FISH as a complement to the conventional cytogenetic studies is of great help in attaining a better characterisation of the chromosome anomalies present in haematological malignancies, such as chromosome 7 monosomy. A study was carried out in three cases of acute non-lymphoblastic leukaemia and a myelodysplastic syndrome with chromosome 7 involvement, as shown by conventional cytogenetic studies. The Cocktail probe for chromosome 7 (DZ1, DZ2) was used (Oncor) in performing in situ hybridation. A monosomic cell line for chromosome 7, undetected by conventional techniques, was disclosed with this procedure in two of the cases. In the remaining two patients the monosomy of chromosome 7 was confirmed, although at percentages different from those attained with the conventional methods.
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