Role of leukemia inhibitory factor during mammalian development.

Leukemia inhibitory factor (LIF) is a cytokine that exhibits proliferative, survival and differentiation activities on a wide range of cell types. A role for LIF in embryonic development is suggested by: i) its ability to stimulate the proliferation of embryonic stem (ES) cells in vitro, while maintaining their totipotency and ii) by both its maternal and embryonic expression at the time of blastocyst implantation. Functional studies of LIF and its receptor during mouse embryogenesis have been performed using the techniques of targeted gene replacement and transgene expression in ES cells to produce transgenic mice bearing either loss- or gain-of-function mutations for LIF activity. Whereas, the phenotype observed in the LIF gain-of-function mutant mice supports a role for LIF in early embryogenesis, the loss-of-function phenotypes point to more specialized functions for LIF in development and further reveal the redundant feature of the LIF cytokine/receptor family.

Leukemia inhibitory factor mediates an injury response but not a target-directed developmental transmitter switch in sympathetic neurons.

Leukemia inhibitory factor (LIF; also known as cholinergic differentiation factor) is a multifunctional cytokine that affects neurons, as well as many other cell types. To examine its neuronal functions in vivo, we have used LIF-deficient mice. In culture, LIF alters the transmitter phenotype of sympathetic neurons, inducing cholinergic function, reducing noradrenergic function, and altering neuropeptide expression. In vivo, a noradrenergic to cholinergic switch occurs in the developing sweat gland innervation, and changes in neuropeptide phenotype occur in axotomized adult ganglia. We find that the gland innervation of LIF-deficient mice is indistinguishable from normal. In contrast, neuropeptide induction in ganglia cultured as explants or axotomized in situ is significantly suppressed in LIF-deficient mice. Thus, LIF plays a role in transmitter changes induced by axotomy but not by developmental interactions with sweat glands.

Leukaemia inhibitory factor is necessary for maintenance of haematopoietic stem cells and thymocyte stimulation.

Leukaemia inhibitory factor (LIF) has a variety of effects on different cell types in vitro, inhibiting the differentiation of embryonic stem cells and promoting the survival and/or proliferation of primitive haematopoietic precursors and primordial germ cells. Here we show that LIF-deficient mice derived by gene targeting techniques have dramatically decreased numbers of stem cells in spleen and bone marrow. Injection of spleen and marrow cells from these mice promotes long-term survival of lethally irradiated wild-type animals, however, showing that the LIF- stem cells remain pluripotent. The numbers of committed progenitors are also reduced in the spleen but not the bone marrow, suggesting that stem cells interact differently with the splenic and medullary microenvironment. Heterozygous animals are intermediate in phenotype, implying that LIF has a dosage effect, and defects in stem cell number can be compensated by exogenous LIF. LIF thus appears to be required for the survival of the normal pool of stem cells, but not their terminal differentiation.

Human monoclonal IgM with autoantibody activity against intermediate filaments.

Monoclonal IgMs from two patients with Waldenström macroglobulinemia were found to react with intermediate filaments. This was shown by (a) immunostaining of various tissues and cultured cells and (b) immunological characterization of the reactive antigen after blotting of polypeptides separated from total cell extracts by gel electrophoresis or purified intermediate filaments on nitrocellulose sheets. One monoclonal IgM had an activity directed only against vimentin, whereas the other reacted with four different classes of intermediate filaments--vimentin, desmin, glial fibrillary protein, and keratins. All of the reactivity of the latter IgM was absorbed by purified vimentin, suggesting that different classes of proteins of intermediate filaments share common antigenic determinant(s). The significance of such autoantibody activity of human monoclonal IgM is discussed in the light of the startling frequency of IgM anti-intermediate filaments antibodies in various diseases.

Tropomyosin synthesis accompanies formation of actin filaments in embryonal carcinoma cells induced to differentiate by hexamethylene bisacetamide.

Hexamethylene bisacetamide (HMBA) induces in vitro the cytodifferentiation of PCC3/A/1 mouse embryonal carcinoma (EC) cells. In EC cells, actin is associated with surface structures but microfilament bundles are not seen. After 2 days of HMBA treatment, rounded EC cells are converted to flat adhesive ones with a developed cytoskeleton containing actin and tropomyosin. The ratio of actin to total proteins is constant in EC cells and their HMBA derivatives; but a striking difference is observed for one of the newly synthesized proteins (Mr 34,000) identified as tropomyosin. Synthesis of tropomyosin is followed by its association with actin microfilament bundles, as revealed by indirect immunofluorescence microscopy with specific antibodies.

[Expression of ts-3 mutant of polyoma virus in mouse cells (author's transl)]. / Expression du mutant ts-3 du virus du polyome dans les cellules de souris.

A thermosensitive mutant (ts-3) of polyoma virus produces T antigen, viral DNA and V antigen in BalbC/3T3 cells only at the permissive temperature (31 degrees C). At the non-permissive temperature (39 degrees C) these cells are unable to complement the viral defect. The pulse technique reveals only transitory cellular DNA induction between 15 and 25 h after infection at 39 degrees C. However, at both temperatures, cytoplasmic penetration is observed, and the viral DNA reaches the nuclei. Similarly, the myoblast cell line PCD2 does not complement the viral defect. In other idfferent types of cells tested (3T6, mouse kidney or secondary embryo cultures), the ts-3 mutant is expressed equally at 31 degrees C and 39 degrees C.

Immunochemical procedure for partial purification of newly synthesized proteins in polyoma virus-infected mouse cells.

A broad-spectrum antiserum has been obtained against nuclear proteins from normal mouse fibroblasts. Solid phase immunoadsorbants prepared from the serum specifically bind the corresponding antigens. The removal in this way of a significant fraction of host proteins leads to partial purification of viral and virus-induced polypeptides from polyoma virus-infected cell extracts. Four main peptides can be selected, with respective molecular weights of 90,000, 70,000, 46,000, and 41,000.