SPECIFIC BINDING OF NERVE GROWTH FACTOR (NGF) BY MURINE C 1300 NEUROBLASTOMA CELLS

Murine C 1300 neuroblastoma cells bind with high avidity on their membrane surface the nerve growth factor (NGF), a protein capable of inducing differentiation of sympathetic nerve cells. The total binding capacity of NGF by the cells was quantitatively measured by a radioimmunoassay technique, using 125I-labeled NGF. An average number of about 106 molecules of NGF could be bound, at saturation, by each cell with an average relative association constant of about 107 liters/mol. Using synchronized cells, it was found, however, that either the number of molecules of ligand bound or the avidity of the binding interaction between NGF and cells varied depending upon their growth cycle, the maximal-binding occurring during the G1 and early S phase. Binding of [125I]NGF was suppressed by trypsin treatment of the cells, however new receptor sites were rapidly replaced onto the membrane surface within 1–2 h. Cells exposed to 3 M KCl released into the supernate a protein product exhibiting similar high avidity for NGF. Acrylamide gel electrophoresis suggested a restricted molecular heterogeneity of this product, with a major component in the 52,000 mol wt region. Antibodies made specific to this protein were capable, in the absence of the complement, of inhibiting the binding of [125I]NGF by the cells and in the presence of the complement they killed them

Ultrastructural studies of spontaneous in vitro transformation of cultured marrow monocyte-macrophage cells from a patient with congenital hypoplastic anemia

The CM-S cell line was established from the bone marrow of a patient suffering from congenital hypoplastic anemia (syndrome of Diamond-Blackfan). The cells grew in suspension in liquid culture and were dependent for their continuous replication in vitro on growth factors produced by the same cells seeded at high density. Initially, undifferentiated blasts, immature myeloid, megakaryocytic and, rarely, erythroid cells were observed. Eventually, a population of cells with characteristics of monocyte-macrophage precursors predominated. These cells could be induced to terminal macrophage differentiation by incubation with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate. During this period (over 150 continuous passages), the cells failed to form colonies in agar and to give rise to tumors when inoculated into athymic mice. On prolonged passages, however, the cells gradually increased their growth capacity in liquid culture and became capable of forming colonies in agar and tumors in animals. Ultrastructural studies revealed that the expression of differentiated traits markedly changed as a function of time: after 277 passages, the transformed cells, although displaying characteristics of monocyte precursors, appeared blocked at this stage and no longer responded to 12-O-tetradecanoylphorbol-13-acetate

Ultrastructural studies of spontaneous in vitro transformation of cultured marrow monocyte-macrophage cells from a patient with congenital hypoplastic anemia

The CM-S cell line was established from the bone marrow of a patient suffering from congenital hypoplastic anemia (syndrome of Diamond-Blackfan). The cells grew in suspension in liquid culture and were dependent for their continuous replication in vitro on growth factors produced by the same cells seeded at high density. Initially, undifferentiated blasts, immature myeloid, megakaryocytic and, rarely, erythroid cells were observed. Eventually, a population of cells with characteristics of monocyte-macrophage precursors predominated. These cells could be induced to terminal macrophage differentiation by incubation with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate. During this period (over 150 continuous passages), the cells failed to form colonies in agar and to give rise to tumors when inoculated into athymic mice. On prolonged passages, however, the cells gradually increased their growth capacity in liquid culture and became capable of forming colonies in agar and tumors in animals. Ultrastructural studies revealed that the expression of differentiated traits markedly changed as a function of time: after 277 passages, the transformed cells, although displaying characteristics of monocyte precursors, appeared blocked at this stage and no longer responded to 12-O-tetradecanoylphorbol-13-acetate

Ultrastructural studies of spontaneous in vitro transformation of cultured marrow monocyte-macrophage cells from a patient with congenital hypoplastic anemia

The CM-S cell line was established from the bone marrow of a patient suffering from congenital hypoplastic anemia (syndrome of Diamond-Blackfan). The cells grew in suspension in liquid culture and were dependent for their continuous replication in vitro on growth factors produced by the same cells seeded at high density. Initially, undifferentiated blasts, immature myeloid, megakaryocytic and, rarely, erythroid cells were observed. Eventually, a population of cells with characteristics of monocyte-macrophage precursors predominated. These cells could be induced to terminal macrophage differentiation by incubation with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate. During this period (over 150 continuous passages), the cells failed to form colonies in agar and to give rise to tumors when inoculated into athymic mice. On prolonged passages, however, the cells gradually increased their growth capacity in liquid culture and became capable of forming colonies in agar and tumors in animals. Ultrastructural studies revealed that the expression of differentiated traits markedly changed as a function of time: after 277 passages, the transformed cells, although displaying characteristics of monocyte precursors, appeared blocked at this stage and no longer responded to 12-O-tetradecanoylphorbol-13-acetate

Nerve growth factor release by human synovial fibroblasts prior to and following exposure to tumor necrosis factor-alpha, interleukin-1 beta and cholecystokinin-8: the possible role of NGF in the inflammatory response.

OBJECTIVE: Aim of this study was to investigate the synthesis, release and effects of nerve growth factor (NGF) in human synovial cells isolated from synovial tissue specimen from healthy and osteoarthritis (OA) patients. METHODS: Human synovial fibroblasts cultures were established starting from healthy and osteoarthritis patients. NGF protein levels in the culture medium, NGFmRNA and high-affinity NGF receptor (Tyrosine kinase A: TrkA) expression in the cells were evaluated in basal conditions and after stimulation with pro-inflammatory cytokines or with the neuropeptide cholecystokinin-8 (CCK-8). The effect of NGF supplement to culture medium on cell proliferation, TrkA expression, and tumour necrosis factor-alpha (TNF-alpha) and inducible-nitric oxide synthase (iNOS) production was investigated. RESULTS: Under basal conditions human synovial cells produce and release NGF. Both interleukin-1-beta (IL-1 beta) and TNF-alpha, but not CCK-8 promote NGF synthesis and release from OA cells. TrkA NGF receptors are also expressed in both normal and OA synovial cells. NGF, but not IL-1 beta, TNF-alpha and CCK-8, enhances the expression of TrkA in isolated synovial cells. NGF down-regulates IL-1 beta-induced TNF-alpha and iNOS production by OA synovial fibroblasts. CONCLUSIONS: NGF is produced and released and TrkA receptors are expressed in synovial inflammation. Overexpression of NGF in inflammed joints might be involved in the modulation rather than in the induction of the joint inflammatory response

Metabolic activation of benzo(alpha)pyrene in fetal mouse hepatocyte lines: Induction of DNA adducts and micronuclei

The 32P-postlabelling assay and the micronucleus test have been used to evaluate spontaneous and induced levels of DNA damage in cultured mammalian cells (C6 and C2.8) following treatment with benzo[a]pyrene and [3H]anti-benzo[a]pyrene diolepoxide

Diagnosi prenatale non invasiva: isolamento di eritrociti nucleati fetali da sangue materno mediante micromanipolazione.

CIC Edizioni Internazional

Nerve growth factor: A survey of activity on immune and hematopoietic cells

Nerve growth factor (NGF) is a well characterized molecule required for the survival and differentiation of a variety of cell types both in the peripheral and central nervous system. Numerous studies published in recent years have demonstrated that NGF affects different functional activities of mature immune and hematopoietic cells. Other studies have revealed that hematopoietic progenitor cells from bone marrow, umbilical cord blood and peripheral blood are receptive to the action of NGF and that bone marrow stromal cells produce/respond to NGF during different steps of normal hematopoiesis. Elevated levels of NGF have been found in a number of inflammatory diseases, including those of autoimmune nature and in myeloproliferative pathologies. This review presents these data and discusses the hypothesis of a possible functional role of NGF in immune and hematopoietic disorders