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Biomedical subjects

I L Weissman

Publications and source records attributed to I L Weissman.

At least 19 recordsLinked to original sources
PubMed1992

Prevention of programmed cell death in Caenorhabditis elegans by human bcl-2.

Programmed cell death is a physiological process that eliminates unwanted cells. The bcl-2 gene regulates programmed cell death in mammalian cells, but the way it functions is not known. Expression of the human bcl-2 gene in the nematode Caenorhabditis elegans reduced the number of programmed cell deaths, suggesting that the mechanism of programmed cell death controlled by bcl-2 in humans is the same as that in nematodes.

Animals↗
PubMed1992

Genomic organization of the mouse granzyme A gene. Two mRNAs encode the same mature granzyme A with different leader peptides.

Granzyme A is a serine protease that, together with the other granular components of cytotoxic T lymphocyte (CTL) cells, has been implicated in the cytolysis process. We report here two different messages and the genomic organization of the mouse granzyme A gene. The granzyme A gene is composed of six exons spanning 7 kilobases. Alternative splicing of the second exon results in the two transcripts. The two mRNA species encode the same mature granzyme A protein but with different leader sequences. The first (HF1) encodes a typical leader signal sequence similar to other granzymes, but the second (HF2) putative leader sequence is different and less hydrophobic. Both messages are present in cultured CTL cell lines and in normal lymphoid tissues. They are both induced when CTL cells are activated in vitro or in vivo. Both messages can be translated in vitro, although the HF1 message appears to be much more efficient as a template. The putative 5' promoter region of the HF gene sequenced (500 base pairs of upstream sequences) contains no well defined promoter sequences aside from the TATA box. The results suggest that (a) granzyme A may be produced with putative different leader sequences from two different mRNAs; (b) this may provide a model system for studying alternate splicing and the evolution of a complex enzymatic system in an organelle; and (c) the genomic DNA reported will be useful for studying transcription regulations involved in controlling the specific expression pattern of this gene.

Amino Acid Sequence↗
PubMed1992

Human T-cell development in SCID-hu mice: staphylococcal enterotoxins induce specific clonal deletions, proliferation, and anergy.

SCID-hu mice provide an in vivo model for studying the events of normal intrathymic human T-cell development and differentiation. We injected SCID-hu mice with staphylococcal enterotoxins (SE) and determined their effects on the development and responsiveness of human T-cell populations defined by their expression of CD4 and CD8, and the type of V beta molecule in their T-cell receptors. After single intraperitoneal injections of SEB or SEE, we observed specific effects on thymic T cells expressing a cognate V beta T-cell receptor (TCR) (V beta 12.1 in the case of SEB-treated SCID-hu mice and V beta 8.1 in the case of SEE-treated mice) using both immunohistochemical staining of thymic frozen sections and flow cytometric analyses. An injection of SEB resulted in a 32% decrease in the total percentages of V beta 12.1+ cells in thymic sections after 2 days, with the greatest effect seen in the medulla, without a demonstrable effect on V beta 5.2/5.3+ or V beta 8.1+ cells. Fluorescence-activated cell sorter analysis demonstrated that TCRhi thymocytes expressing a cognate V beta TCR declined transiently by 35% to 45% 1 to 2 days after the injection of SE. Analysis of thymic subpopulations showed decreases in the TCRhi CD4+8- and CD4-8+ cells and an increase in TCRlo CD4-8+ cells. Multiple injections of SE resulted in 50% to 60% decreases in cognate V beta TCR+ CD4+8- populations. Thymocytes prepared from SE-treated SCID-hu mice demonstrated specific anergy to the SE to which they had previously been exposed in vivo, but had a normal proliferative response to other superantigens in an in vitro assay. In contrast to the effects on thymic T cells, single injections of SE resulted in a twofold increase in the total numbers of circulating CD4+8- and CD4-8+ human T cells and a fourfold to eightfold increase in T cells expressing a cognate V beta TCR. Using SE as superantigens in SCID-hu mice, we have been able to induce antigen-specific clonal deletions, anergy, and proliferation of human T cells.

Animals↗
PubMed1992

Cloning and expression of mouse integrin beta p(beta 7): a functional role in Peyer's patch-specific lymphocyte homing.

Lymphocytes express integrin receptors, termed lymphocyte Peyer's patch high endothelial venule (HEV) adhesion molecules (LPAMs), that mediate their organ-specific adhesion to specialized HEVs found in mucosal lymphoid organs (Peyer's patches). LPAM-1 consists of a murine integrin alpha 4 noncovalently associated with integrin beta p. Here, we describe the cloning and expression of a mouse cDNA encoding beta p, which is an 806-amino acid transmembrane glycoprotein. The genomic Southern blot analysis indicates that beta p is the murine homologue of human beta 7. The function of alpha 4 beta 7 as a Peyer's patch-specific adhesion molecule was tested directly by expression of the murine beta 7 cDNA in an alpha 4+ beta 7-B-cell line or coexpression of the alpha 4 and beta 7 cDNAs in an alpha 4-beta 7-T-cell line. The transfected cells exhibited a new Peyer's patch-specific adhesive phenotype that could be specifically blocked by monoclonal antibodies against alpha 4 and beta 7. Moreover, an anti-beta 7 monoclonal antibody specifically blocked binding of normal lymphocytes to Peyer's patch HEV but did not inhibit their binding to peripheral lymph node HEVs, indicating that beta 7 is a unique component of the Peyer's patch-specific homing receptor.

Amino Acid Sequence↗
PubMed1992

Incidents of rejection and indifference in Fu/HC incompatible protochordate colonies.

We test here for the existence of specific alloimmune memory in the rejection responses of the colonial tunicate Botryllus schlosseri. Colony specificity in these organisms is controlled by the Fu/HC locus. Rejection occurs only between colonies that do not share any allelic determinant at this locus. Two sets of experiments were conducted: 1) Forty-nine pairs of nonfusible oozooids were interacted naturally along a period of several months. They rejected, disconnected, and in 20% of the cases, interacted again. 2) Repeated colony allorecognition assays were done on 15 pairs of interacting subclones (up to 5 consecutive tests/pair). Major results indicate: 1) Not all ampulla-ampulla interactions resulted in necrotic areas, points of rejection (PORs). 2) A full repertoire of PORs was attained within the first 10 days. Thereafter, no more PORs were added. 3) The outcome of indifference (cases where ampulla-ampulla contacts did not result in rejection) was repeatedly recorded in multiple tests, and its frequency increased in the secondary and tertiary tests along a set of 5 consecutive tests. It is concluded that allospecific memory, as measured by an accelerated production of PORs and amplification in their number, was not characteristic of the Botryllus rejection phenomenon, which, however, reveals the qualities of a low responder. These results are discussed in the light of some aspects of tolerance in the mammalian system.

Animals↗
PubMed1992

Unexpected effects of the severe combined immunodeficiency mutation on murine lymphomagenesis.

Strain C.B17 scid/scid (SCID) mice, which lack functional T and B lymphocytes, show heightened susceptibility to the induction of thymic lymphomas by x-irradiation. Susceptibility is highest in thymus-chimeric SCID-BL mice (thymectomized SCID mice bearing a C57BL thymus graft). All SCID-BL lymphomas originate in the cells of the thymic graft (C57BL type) and lack murine leukemia virus expression. Both SCID and SCID-BL lymphomas are phenotypically CD4-8+ and/or CD4+8+, but only the SCID-BL tumors express CD3. Injection of C57BL or BALB/c bone marrow into irradiated SCID-BL mice prevents lymphoma development, but SCID marrow is completely ineffective. The results suggest that the scid condition enhances the activity of a putative lymphomagenic agent induced in the bone marrow by x-irradiation and that C57BL thymic cells are highly sensitive targets. Moreover, the failure of SCID bone marrow to protect against lymphomagenesis vs. the efficacy of marrow from immunocompetent donors points to involvement of T or B lineage cells in this process.

Animals↗
PubMed1992

Development of gamma delta T-cell subsets from fetal hematopoietic stem cells.

Hematopoietic stem cells (HSC) were isolated from mouse fetus, and their developmental potential was compared with adult HSC. Donor-derived V gamma 3+T cells were detected in fetal thymic lobes, repopulated in vitro with fetal liver HSC, but not in those with adult bone marrow HSC. Single clonogenic fetal HSC gave rise to thymic progeny that include V gamma 3+, other gamma delta+, and alpha beta+ T cells. No V gamma 3+ T cells were detected in adult thymus injected intrathymically with either fetal or adult HSC. These results support a hypothesis that only fetal HSC have the capacity to differentiate into V gamma 3+ T cells in the fetal thymic microenvironment, and that the developmental potential of HSC may change during ontogeny.

Aging↗
PubMed1992

Evidence for a programmed life span in a colonial protochordate.

The variety of theories that have attempted to define the mechanisms of aging and life span can be broadly divided into two alternative but nonexclusive viewpoints. The first stipulates that random changes of cellular and molecular structures lead to death following progressive "wear and tear." The second argues that life span is, at least in part, genetically programmed, and therefore aging may also result from time-dependent intrinsic processes. Here we demonstrate that ramets (clonal replicates) experimentally separated from colonies of the acidian protochordate Botryllus schlosseri died months after their separation, almost simultaneously with their parent colony and sibling ramets. In addition, in experimentally joined chimeras between ramets of senescent and nonsenescent colonies, elements from different parent colonies displayed parent-colony-specific timing of mortality. Thus, the senescent phenotype was simultaneously expressed both in chimeras and in unfused ramets of the parent colony that was undergoing senescence, whereas control ramets from the other partner survived. These findings provide experimental evidence for a heritable basis underlying mortality in protochordates, unlinked to reproductive effort and other life history traits of this species.

Aging↗
PubMed1992

Mouse MRP8 and MRP14, two intracellular calcium-binding proteins associated with the development of the myeloid lineage.

MRP8 and MRP14 are two S100-like calcium-binding proteins of unknown function, associated with numbers of human inflammatory disorders. Both molecules have been described as L1 complex, cystic fibrosis antigen, or p8 and p14. We report here the cloning of mouse MRP8 and MRP14 and their pattern of expression during hematopoiesis. Mouse MRP8 and MRP14 proteins share 59% identity with their human counterparts, but they are more divergent than the other members of the S100 protein family. Mouse MRP proteins are coexpressed in fetal myeloid progenitors, where they are detected as early as day 11 of gestation. In fetal liver and yolk sac, MRP+ cell populations increased in number, in association with the development of the myeloid lineage. In adult mouse, we identified MRP8 and MRP14 proteins in immature myeloid cells of the bone marrow, myeloid cells in the splenic red pulp and marginal zone, in addition to monocytes and blood neutrophils. However, MRP expression is lost as cells terminally differentiate into tissue macrophages. In addition, using thioglycollate-induced peritoneal inflammatory exudates, we showed that MRP8 and MRP14 proteins are highly expressed in recruited neutrophils and monocytes.

Animals↗
PubMed1992

Isolation of a candidate human hematopoietic stem-cell population.

We have identified a rare (0.05-0.1%) subset of human fetal bone marrow cells that contains multipotent hematopoietic precursors. The population of human precursor cells that express Thy-1 and CD34 but no known lineage markers is enriched for clonogenic activity that establishes long-term, multilineage (myelomonocytic and B lymphoid) cultures on mouse marrow stromal lines. Further, the Thy-1+CD34+ subset that takes up little of the fluorescent mitochondrial dye rhodamine 123 contains virtually all the cells that establish long-term cultures. In human fetal thymus transplanted into SCID (severe combined immunodeficiency) mice, Thy-1+CD34+ fetal bone marrow cells differentiate into T lymphocytes. In two of nine cases, allogeneic Thy-1+CD34+ cells could engraft intact human fetal bone marrow grown in SCID mice, resulting in donor-derived myeloid and B cells. By extrapolation, the rare human Thy-1+Lin-CD34+ cell population contains pluripotent hematopoietic progenitors; we propose that it is highly enriched for candidate hematopoietic stem cells.

Animals↗
PubMed1992

Evidence that hematopoietic stem cells express mouse c-kit but do not depend on steel factor for their generation.

The interaction of the mouse c-kit receptor, designated Kit receptor, and steel factor promotes the proliferation and differentiation of hematopoietic progenitor cells. Monoclonal antibodies against the extracellular portion of the mouse Kit receptor were established. Five percent to 10% of total bone marrow cells expressed the Kit receptor, and half of them lack the expression of lineage markers. The Kit receptor was expressed on 70-80% of Thy-1.1lo Lin-Sca-1+ cells, which express Thy-1.1 antigen at a low level and constitute approximately 0.05% of adult bone marrow and fetal liver; by previous studies, these cells have been shown to be highly enriched for multipotent hematopoietic stem cells (HSCs) and are the only hematopoietic cell subset with this activity. Spleen colony formation and long-term multilineage reconstitution activities were contained in the Kit+ but not in the Kit- subpopulations of Thy-1lo Lin-Sca-1+ cells from adult bone marrow, suggesting that the Kit receptor is expressed on HSCs from the earliest stage-i.e., pluripotent HSCs. The role of steel factor in the development and self-renewal of HSCs was tested with Sl/Sl homozygote fetuses, which lack genes to encode functional steel factor. They were shown to have 30-40% of the number of HSCs on days 13-15 when compared with normal litermates. However, the absolute number of HSCs increased during fetal development in the Sl/Sl mice. The results suggest that the Kit receptor-steel factor interaction may not be essential for the initiation of hematopoiesis and the self-renewal of (at least) fetal HSCs.

Animals↗
PubMed1992

Perforin and granzyme A expression identifying cytolytic lymphocytes in rheumatoid arthritis.

Lymphocytes from the synovial fluid of patients with rheumatoid arthritis were examined for the expression of granzyme A and perforin. Previous studies have demonstrated that the expression of these proteins, which are implicated as mediators of cytotoxicity, can be used to identify putative cytolytic lymphocytes in vivo. Twenty-two synovial fluid samples were analyzed by in situ hybridization and immunohistochemistry. In six patients receiving low doses of immunosuppressant, a population of granzyme A- and perforin-expressing lymphocytes could be identified. In contrast, lymphocytes from patients who were receiving high doses of immunosuppressant did not contain any granzyme A- or perforin-expressing lymphocytes. Synovial fluid lymphocytes from patients with osteoarthritis did not express either marker. The expression of these markers demonstrates the presence of potentially functional cytolytic lymphocytes, expressing proteins required to mediate killing, in the synovial fluid of patients with rheumatoid arthritis. This suggests that cytolytic lymphocytes may be involved in the pathogenesis of rheumatoid arthritis.

Arthritis, Rheumatoid↗
PubMed1992

Searching for hematopoietic stem cells: evidence that Thy-1.1lo Lin- Sca-1+ cells are the only stem cells in C57BL/Ka-Thy-1.1 bone marrow.

Hematopoietic stem cells (HSCs) are defined in mice by three activities: they must rescue lethally irradiated mice (radioprotection), they must self-renew, and they must restore all blood cell lineages permanently. We initially demonstrated that HSCs were contained in a rare (approximately 0.05%) subset of bone marrow cells with the following surface marker profile: Thy-1.1lo Lin- Sca-1+. These cells were capable of long-term, multi-lineage reconstitution and radioprotection of lethally irradiated mice with an enrichment that mirrors their representation in bone marrow, namely, 1,000-2,000-fold. However, the experiments reported did not exclude the possibility that stem cell activity may also reside in populations that are Thy-1.1-, Sca-1-, or Lin+. In this article stem cell activity was determined by measuring: (a) radioprotection provided by sorted cells; (b) long-term, multi-lineage reconstitution of these surviving mice; and (c) long-term, multi-lineage reconstitution by donor cells when radioprotection is provided by coinjection of congenic host bone marrow cells. Here we demonstrate that HSC activity was detected in Thy-1.1+, Sca-1+, and Lin- fractions, but not Thy-1.1-, Sca-1-, or Lin+ bone marrow cells. We conclude that Thy-1.1lo Lin- Sca-1+ cells comprise the only adult C57BL/Ka-Thy-1.1 mouse bone marrow subset that contains pluripotent HSCs.

Animals↗
PubMed1992

A cyclical, developmentally-regulated death phenomenon in a colonial urochordate.

Botryllus schlosseri is a colonial ascidian whose asexually derived, clonally modular systems of zooids exhibit developmental synchrony. The blastogenic cycle culminates in a phase of programmed cell and zooid death called takeover, in which all functional zooids die over a 30 hr period, and are replaced by a new generation of individuals. Because of the weekly recurrence and magnitude of visceral death in this model organism, we have begun to characterize the mechanisms that govern takeover. Here we describe a monoclonal antibody (B3F12.9) that recognizes a novel 57 Kd polypeptide (under reducing conditions) localized to the perivisceral extracellular matrix (PVEM) of buds and zooids, as well as blood cells of Botryllus by immunofluorescence and immunogold labeling of tissue sections. During their active feeding phase, zooids exhibited a uniform labeling pattern of PVEM along their anteroposterior (A-P) axis. At the onset of takeover (T = 3 hr), B3F12.9 immunostaining became diffuse or absent at the anterior end, which paralleled the axis of contraction of the dying zooid, whereas the posterior end retained its labeling integrity. During mid (T = 15 hr) to late (T = 28 hr) takeover, issue damage was extensive, large blood macrophages and other B3F12.9 immunoreactive blood cells invaded the peribranchial cavity, whereas PVEM labeling gradually disappeared along the entire A-P axis. These findings indicate that takeover is a dynamic process in which extracellular matrix breakdown proceeds in a polarized fashion, beginning at the anterior end of each zooid and gradually propagating toward the posterior end.

Animals↗
PubMed1992

Bcl-2 prevents death of factor-deprived cells but fails to prevent apoptosis in targets of cell mediated killing.

'Programmed cell death' has been used to describe the death of cells killed by cytotoxic T cells or growth factor deprivation. Although bcl-2 can prevent death of cells deprived of growth factor, it failed to protect cells against T cell killing. In spite of bcl-2 expression, the DNA of targeted cells was degraded into nucleosome-sized fragments. Therefore the early steps in apoptosis induced by factor deprivation differ from those triggered by cytotoxic T cells, although they share a common final pathway featuring degradation of the DNA and loss of cytoplasmic membrane integrity.

Animals↗
PubMed1992

Lymphocyte development from stem cells.

Highly enriched pluripotent and multipotent hematopoietic stem cells (HSCs) are isolated from bone marrow and fetal liver as Thy-1loLin-Sca-1+ cells. Pluripotent HSCs express c-kit receptor on their surface, but the generation and proliferation of early fetal HSCs take place in the absence of steel factor. T precursor cells migrate into the fetal thymus by chemotactic mechanism. CD4lo precursors represent a newly defined phase of T-cell development in the thymus between the bone marrow-derived stem cells and the CD4-8- intrathymic precursors. Only fetal, but not adult, HSCs have the capacity to differentiate into V gamma 3+ and V gamma 4+ T cells under the fetal thymic microenvironment, and HSC themselves may lose some of their developmental potential during ontogeny. It is postulated that HSCs are the locus of a complicated but precise developmental clock that may determine both the time-dependent closure of some gene loci (e.g. V gamma 3 and V gamma 4 T cell receptor, and embryonic and fetal globin) and the activation of others (e.g. the N nucleotide insertion machinery).

Animals↗