Biomedical subjects

L Tessarollo

Publications and source records attributed to L Tessarollo.

49 records · Page 3Linked to original sources

PubMed1995

Neurotrophin and neurotrophin receptors in vascular smooth muscle cells. Regulation of expression in response to injury.

The neurotrophins, a family of related polypeptide growth factors including nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and neurotrophin (NT)-3 and NT-4/5 promote the survival and differentiation of distinctive sets of embryonic neurons. Here we define a new functional role for neurotrophins, as autocrine or local paracrine mediators of vascular smooth muscle cell migration. We have identified neurotrophins, and their cognate receptors, the trk tyrosine kinases, in human and rat vascular smooth muscle cells in vivo. In vitro, cultured human smooth muscle cells express BDNF; NT-3; and trk A, B, and C. Similarly, rat smooth muscle cells expressed all three trk receptors as well as all four neurotrophins. Moreover, NGF induces cultured human smooth muscle cell migration at subnanomolar concentrations. In the rat aortic balloon deendothelialization model of vascular injury, the expression of NGF, BNDF, and their receptors trk A and trk B increased dramatically in the area of injury within 3 days and persisted during the formation of the neointima. In human coronary atherosclerotic lesions, BDNF, NT-3, and NT-4/5, and the trk B and trk C receptors could be demonstrated in smooth muscle cells. These findings suggest that neurotrophins play an important role in regulating the response of vascular smooth muscle cells to injury.

Animals↗

PubMed1994

Targeted mutation in the neurotrophin-3 gene results in loss of muscle sensory neurons.

Neurotrophin 3 (NT-3) is one of four related polypeptide growth factors that share structural and functional homology to nerve growth factor (NGF). NT-3 and its receptor, called neurotrophic tyrosine kinase receptor type 3 (Ntrk3; also called TrkC), are expressed early and throughout embryogenesis. We have inactivated the NT-3 gene in embryonic stem (ES) cells by homologous recombination. The mutated allele has been transmitted through the mouse germ line, and heterozygote intercrosses have yielded homozygous mutant newborn pups. The NT-3-deficient mutants fail to thrive and exhibit severe neurological dysfunction. Analysis of mutant embryos uncovers loss of Ntrk3/TrkC-expressing sensory neurons and abnormalities at early stages of sensory neuronal development. NT-3-deficient mice will permit further study of the role of this neurotrophin in neural development.

Animals↗

PubMed1993

The rat trkC locus encodes multiple neurogenic receptors that exhibit differential response to neurotrophin-3 in PC12 cells.

Members of the Trk tyrosine kinase family have recently been identified as functional receptors of the NGF family of neurotrophins. Here we show the rat trkC locus to be complex, encoding at least four distinct polypeptides. Three of the encoded polypeptides are full-length receptor tyrosine kinases that differ by novel amino acid insertions in the kinase domain. A fourth protein is a truncated receptor that lacks the catalytic domain. Tyrosine phosphorylation, cross-linking, and ligand binding assays indicate that TrkC receptors interact with NT-3 and not with the related neurotrophins NGF, BDNF, xNT-4, or hNT-5. Furthermore, high and low affinity NT-3-binding sites are associated with the TrkC receptors. Stable and transient expression of TrkC receptors in PC12 cells indicates that the neurite outgrowth response elicited by NT-3 is dramatic in receptors lacking the novel kinase insert (gp150trkC) but absent in receptors containing the 14 amino acid insert in the kinase domain (gp150trkC14). These data suggest that the trkC locus encodes receptors that may be capable of mediating different biological responses within the cell. This could have important implications in understanding the role of neurotrophins in the development of the vertebrate nervous system.

3T3 Cells↗

PubMed1993

Neurotrophin-3 induced by tri-iodothyronine in cerebellar granule cells promotes Purkinje cell differentiation.

Thyroid hormones play an important role in brain development, but the mechanism(s) by which triiodothyronine (T3) mediates neuronal differentiation is poorly understood. Here we demonstrate that T3 regulates the neurotrophic factor, neurotrophin-3 (NT-3), in developing rat cerebellar granule cells both in cell culture and in vivo. In situ hybridization experiments showed that developing Purkinje cells do not express NT-3 mRNA but do express trkC, the putative neuronal receptor for NT-3. Addition of recombinant NT-3 to cerebellar cultures from embryonic rat brain induces hypertrophy and neurite sprouting of Purkinje cells, and upregulates the mRNA encoding the calcium-binding protein, calbindin-28 kD. The present study demonstrates a novel interaction between cerebellar granule neurons and developing Purkinje cells in which NT-3 induced by T3 in the granule cells promotes Purkinje cell differentiation.

Animals↗

PubMed1993

trkC, a receptor for neurotrophin-3, is widely expressed in the developing nervous system and in non-neuronal tissues.

The Trk family of tyrosine kinases encodes receptors for nerve growth factor-related neurotrophins. Here we present a developmental expression study of trkC, which encodes a receptor for neurotrophin-3 (NT-3). Like the related genes, trk and trkB, trkC is expressed primarily in neural lineages although the pattern is complex and includes non-neuronal cells. Direct comparison with trk and trkB developmental expression patterns permits the following observations. (1) trkC is expressed in novel neural tissues where other Trk genes are silent. (2) Some tissues appear to coexpress trkB and trkC receptors in the embryo and in the adult. (3) trkC expression can be detected in the gastrulating embryo. These data provide insights into the role of Trk-family receptors and nerve growth factor-related neurotrophins during development and suggest that, in addition to regulating neuronal survival and differentiation, the neurotrophin/Trk receptor system may have broader physiological effects. Finally, interspecific mouse backcrosses have been used to map the location of each of the Trk genes on mouse chromosomes. Alignment with available chromosomal maps identify possible linkage between the Trk genes and known neurological mutations.

Animals↗

PubMed1992

Tumours induced by Moloney murine sarcoma virus are clonal in rats, not clonal in mice.

Moloney murine sarcoma virus (M-MSV) induces rapidly growing tumours in adult mice of most conventional strains. Rats are less susceptible to M-MSV oncogenesis, but the few rhabdomyosarcomas that do develop after viral inoculation of newborn animals closely resemble conventional malignancies: they develop after a long latency, grow progressively, and metastasize to regional lymph nodes and lungs. Southern blot analysis with a v-mos-specific probe of M-MSV-induced tumours in both species demonstrated an oligo-, monoclonal pattern of exogenous v-mos integration only in the rat system, while mouse tumours were not clonal in origin. Furthermore, the same type of analysis of lymph node and lung metastases showed that cell clones already present in the primary rat lesion colonized secondary sites during tumour progression. Apparently, Moloney murine leukemia virus (M-MuLV) was not involved in rhabdomyosarcoma pathogenesis since M-MuLV-specific DNA sequences could not be demonstrated in three of the six rat tumours. Finally, in all mouse tumours, unintegrated linear M-MSV proviruses could be readily detected.

Animals↗

PubMed1992

In vivo and in vitro death of mature T cells induced by separate signals to CD4 and alpha beta TCR.

To investigate whether a clonal deletion mechanism is responsible for the mature T cell tolerance that may be induced in vivo by TCR signal to anti-CD4 (H129.19 mAb) coated cells, we analyzed the T cell repertoire in anti-CD4 mAb treated BALB/c mice by flow cytometry following TCR signals through anti-alpha beta TCR mAb or SEB superantigen. Lymph nodes showed a strong reduction in the CD4+/CD8+ cell ratio, and a selective clonal loss of CD4+ V beta 8+ cells 4d following anti-alpha beta TCR or SEB injection, respectively. Following lymph node cell activation in a short-term in vitro assay with SEB or anti-V beta 8 mAb, a selective elimination of CD4+ V beta 8+ cells was again detected, and DNA fragmentation analysis disclosed a cell death by apoptosis. These findings suggest that TCR triggering transduces an apoptotic signal into CD4+ mAb saturated cells that in turn leads to specific holes in the mature T cell repertoire.

Animals↗

PubMed1992

Loss of N-myc function results in embryonic lethality and failure of the epithelial component of the embryo to develop.

myc genes are thought to function in the processes of cellular proliferation and differentiation. To gain insight into the role of the N-myc gene during embryogenesis, we examined its expression in embryos during postimplantation development using RNA in situ hybridization. Tissue- and cell-specific patterns of expression unique to N-myc as compared with the related c-myc gene were observed. N-myc transcripts become progressively restricted to specific cell types, primarily to epithelial tissues including those of the developing nervous system and those in developing organs characterized by epithelio-mesenchymal interaction. In contrast, c-myc transcripts were confined to the mesenchymal compartments. These data suggest that c-myc and N-myc proteins may interact with different substrates in performing their function during embryogenesis and suggest further that there are linked regulatory mechanisms for normal expression in the embryo. We have mutated the N-myc locus via homologous recombination in embryonic stem (ES) cells and introduced the mutated allele into the mouse germ line. Live-born heterozygotes are under-represented but appear normal. Homozygous mutant embryos die prenatally at approximately 11.5 days of gestation. Histologic examination of homozygous mutant embryos indicates that several developing organs are affected. These include the central and peripheral nervous systems, mesonephros, lung, and gut. Thus, N-myc function is required during embryogenesis, and the pathology observed is consistent with the normal pattern of N-myc expression. Examination of c-myc expression in mutant embryos indicates the existence of coordinate regulation of myc genes during mouse embryogenesis.

Alleles↗

PubMed1992

The Trk family of tyrosine kinases: receptors for NGF-related neurotrophins.

Neurotrophins are known to have important functions in the survival of embryonic and adult subpopulations of neurons. The identification of Trk family RTKs as receptors for NGF-related neurotrophins indicates phosphotyrosine-mediated signal transduction as a principal mechanism for neurotrophin signaling. Previous trk and trkB expression studies (Klein et al. 1989, 1990b; Martin-Zanca et al. 1990) and more recent studies with trkC (L. Tessarollo et al., in prep.) provide important clues about function. Thus, trkB and trkC expression in motor neurons and in many nonneuronal cells suggests that these cells are targets for neurotrophin action in vivo, even though this has not been demonstrated in the classic in vitro survival assays. Expression of trkB and trkC in nonneuronal cells implies that these receptors may act in additional aspects of organogenesis and development. Current approaches to assay Trk receptor and neurotrophin function will be complemented by further studies in the living organism. Transgenic approaches aimed at ectopic expression and at interfering with normal receptor function should provide additional insights. Finally, reverse genetic approaches using targeted mutation of Trk receptors in embryonic stem cells (Stanton et al. 1992) will allow assessment of critical receptor requirements and provide powerful reagents for studying nervous system development and function.

Animals↗

PubMed1992

c-ros: the vertebrate homolog of the sevenless tyrosine kinase receptor is tightly regulated during organogenesis in mouse embryonic development.

The c-ros proto-oncogene is the vertebrate homologue of the Drosophila sevenless tyrosine kinase receptor. Examination of c-ros mRNA transcripts in the mouse embryo reveals a stringent pattern of expression. Only kidney, intestine and lung exhibit ros-specific RNA using sensitive techniques such as RNAase protection and in situ hybridization. The temporal and spatial arrangement of c-ros transcripts is coincident with the phenotypic induction and proliferation of epithelium during organogenesis of the kidney and intestine. The data provide evidence for a role of c-ros in the obligate cell-cell interactions that characterize the morphogenic induction and proliferation of epithelial cells in the kidney, intestine and lung. The c-ros tyrosine kinase receptor may provide a signal transduction pathway for epithelial-mesenchymal interactions.

Animals↗

PubMed1991

An improved method for the detection of DNA fragmentation.

An application of the Southern blot technique is described which permits the detection of DNA fragmentation due to cell death by apoptosis. DNA fragments were isolated from cell suspensions and tissues, separated on agarose gel, transferred by Southern blot and hybridized with a radiolabeled total cellular DNA probe. The application of this procedure to thymus cell samples, revealed the distinct ladder pattern of DNA fragments in multiples of about 180-200 base pairs, a characteristic feature of DNA fragmentation. In comparison to conventional DNA visualization with ethidium bromide staining, the radiolabeled probe improved the detection of DNA fragments at least eight-fold. This method detects low levels of DNA fragments, as well as physiological tissue DNA fragmentation, while avoiding cell damage due to DNA radiolabeling.

Animals↗

PubMed1991

In vitro differentiation of rhabdomyosarcomas induced by nickel or by Moloney murine sarcoma virus.

In vitro cultures and clonal derivatives have been established from rat rhabdomyosarcomas induced by Moloney-Murine Sarcoma Virus (MSV) or by nickel sulfide; differentiation ability has been studied as expression of desmin, embryonic and adult myosin isoforms, alpha-actin isoforms and cellular fusion. The two rhabdomyosarcoma models showed different levels of myogenic differentiation. Multinucleated myotube-like structures were frequently observed in cultures derived from nickel-induced tumours. Desmin was present in 50-80% of cells and embryonic myosin in up to 10%. In MSV-tumour-derived cultures and in their metastases or clonal derivatives two cell types are present in different ratios: spindle-shaped cells, adherent to plastic surfaces, and rounded cells, loosely attached or floating free in the medium. These cultures showed features of myogenic differentiation (10-80% desmin-positive cells), but embryonic myosin expression and production of multinucleated myotube-like structures were very rare events. Cultures from autochthonous lymph node and lung metastatic cells showed similar patterns of differentiation. Retinoic acid increased differentiated features (myotube formation and embryonic myosin expression) only in nickel-induced rhabdomyosarcoma cells. The two models described here mimic the heterogeneity in differentiation pattern found among human rhabdomyosarcomas. Myogenic differentiation ability was retained at a good level by nickel-induced tumours, whereas it was strongly impaired in MSV-induced tumours.

Animals↗

PubMed1989

Human monoclonal antibody against a gag-coded protein of human immunodeficiency virus produced by a stable EBV-transformed cell clone.

An EBV-transformed lymphoblastoid B cell clone (A12) derived from peripheral blood lymphocytes of an HIV-1-infected individual is described. The immunoglobulin isotype produced by this clone was IgM, and Southern blot analysis of immunoglobulin gene rearrangement showed a monoclonal pattern. The A12 monoclonal antibody was specific for the p24 product of the HIV-1 gag gene. This clone is now in continuous culture for more than 8 months and no changes in its biologic properties have been observed.

Antibodies, Monoclonal↗