In vitro duplication and in vivo cure of mast-cell deficiency of Sl/Sld mutant mice by cloned 3T3 fibroblasts.

Abstract

S1/Sd mutant mice are profoundly deficient in tissue mast cells as a result of a defect in the microenvironment promoting the development ofthese cells. To facilitate the analysis of the SI mutation, we attempted to establish an in vitro system in which the in vivo defect of S/1Sd mice could be A homogenous population of mast cells has been generated in vitro by cultivating progenitors from bone marrow in the presence of a T-cell-derived growth factor, interleukin (IL) 3 (for reviews, see refs. 7 and 8). The action of IL-3 has been shown to be enhanced by another T-cell-derived growth factor, IL-4 (9, 10). Recently we described another mode of mast-cell growth, which depends on contact with fibroblasts (11). When mast cells obtained in vitro from bone-marrow cells were cocultured with the NIH 3T3 fibroblast cell line, the mast cells were maintained in the absence of an exogenous source of IL-3 and IL-4. We further demonstrated that cultured mast cells derived from W/WV mice were defective in fibroblast-dependent growth, although they responded to T-cell-derived growth factors as efficiently as those from +/+ mice (12).In the present study we examined whether the mast-cell deficiency in 51/51d mice could be reproduced in vitro by a similar coculture system. No abnormality in proliferative response to either T-cell-derived Nakahata et al. (13). Homogenous populations of mast cells were obtained by culturing bone-marrow cells at 106 cells per ml in a-minimal essential medium supplemented with 10-4 M 2-mercaptoethanol/20% (vol/vol) horse serum/10% (vol/vol) PWM-SCM. Half of the medium was replaced every 7 days. Four weeks after initiation of the culture, >98% of the cultured cells were identified as mast cells, and 13% of them were found to be in S phase as described (11). Therefore, those cultured for >4 wk were used in these experiments. The NIH 3T3 fibroblast cell line derived from a Swiss mouse embryo (14) or other fibroblast cell lines were cocultured with mast cells in the absence of PWM-SCM as described (11,12). Briefly, 105 cultured mast cells suspended in 2 ml of medium containing 5% fetal calf serum but not PWM-SCM were added to a confluent culture Abbreviations: IL, interleukin; PWM-SCM, pokeweed mitogenstimulated spleen cell-conditioned medium; BrdUrd, 5-bromodeoxyuridine. *To whom reprint requests should be addressed. 2888The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

Transformation of NIH3T3 fibroblasts by the c-Kit receptor tyrosine kinase: effect of receptor density and ligand-requirement

“…In order to investigate the transforming potential of c-Kit in the absence of endogenous SLF production, Sl/Sl-3T3 cells (Fujita et al, 1989), which lack SLF mRNA (c.f. Caruana et al, 1993) were infected with the Zen(mukit) retroviral construct.…”

Co expression of SCF and KIT in gastrointestinal stromal tumours (GISTs) suggests an autocrine/paracrine mechanism

“…SCF exists under soluble (s) and membranebound (m) forms due to differential splicing and proteolytic cleavage (Broudy, 1997). The two forms display distinct effects as regards to the survival and proliferation of haematopoietic cell lines (Caruana et al, 1993;Miyazawa et al, 1995) and primary cells (Fujita et al, 1989;Toksoz et al, 1992), although they are both active in increasing the number of human progenitor cells in the context of stromal cell cultures (Toksoz et al, 1992). Stromal mSCF appears to induce more persistent signalling than the soluble form, this last form inducing rapid downregulation of cell surface expression and degradation of KIT (Miyazawa et al, 1995).…”

The Role of Cytokines in Human Endometrium: The Inhibitory Effect of IL-1 and TNF α on <I>in Vitro</I> Decidualization and mRNA Expression of M-CSF, SCF and LIF in the Human Endometrium

“…In culture experiments with RT-PCR,SCF mRNA expression was much stronger in the stromal cell-enriched fraction(SF)than in the epithelial glandular cell-enriched fraction(EF) (Fig.9).By contrast even after 9 days culture with progesterone or estrogen,no change in SCF mRNA expression was seen when compared to the control culture without steroids(data not shown).These findings suggest that SCF in non-pregnant endometrium [31,32].In mice,the c-kit protein was detected in the uterus,the decidua basalis facing the invading trophoblasts,and the placental labyrinthine layer during pregnancy [6,7].Therefore,endogenous SCF in mouse uterus is supposed to play an important role in early embryonic development in the peri-implantation period.…”